ExperimentHPLC
ExperimentHPLC[Samples]⟹Protocol
generates a Protocol to separate Samples via high-pressure liquid chromatography (HPLC).
High-performance liquid chromatography (HPLC) involves an aqueous and/or organic mobile phase, into which, samples are injected and flowed through the stationary phase column. Analytes pass over resin within the column and selectively adsorb based on their relative affinity, leading to differential retention of unique analyte molecules. Molecules are carried downstream for chemical property analysis and optional collection. On the whole, molecular components are separated into analyzable, collectable constituents. For instance, synthesized DNA can be purified based on the total ionic charge of each unique strand.
Experimental Principles
Figure 1.1: Procedural overview of a HPLC experiment. Step 1: If ConductivityCalibration -> True, the conductance probe is calibrated and the conductance flow cell is rinsed with cleaning solutions. Step 2: If pHCalibration -> True, the pH probe is calibrated and the pH flow cell is rinsed with cleaning solutions. Step 3: During the system prime, the system is rinsed with cleaning solutions. Step 4: The stationary Column is installed and equilibrated to measurement conditions. Step 5: Samples are then introduced into the flow path. Step 6: The analytes are selectively retained on the downstream Column. Step 7: Upon exit from the column, the now separated analytes are analyzed according to their physical property. Step 8: Based on the property measurement and specifications, analytes are saved in output containers, if CollectFractions ->True. Step 9: After final sample measurement, the Column is rinsed, removed from the system, and stowed. Step 10: After the column is removed the system is rinsed with cleaning solution.
Instrumentation
Waters Acquity UPLC H-Class PDA
Figure 2.1.1: Instrument diagram for the H-Class and I-Class systems. Buffer solutions (up to 4) are steadily pumped through the instrument, consisting of a 6-port valve system, adsorbent column, detectors, and terminates at fraction collection or waste. Samples within the autosampler are loaded into the sample loop via positive displacement from the syringe. The rotation of the valve exposes the sample loop, thereby carrying the sample downstream to the column. Within the column, molecular constituents are separated by adsorption -- a function of the buffers, column, sample properties and temperature. Analytes are continuously carried downstream to the detectors, where they can elicit quantifiable peak signals.
Figure 2.1.2: Principle of PhotodiodeArray detection. Filtered light (across a range of wavelengths) passes through the flow downstream of the column. Presence of light-absorbing molecules will result in less light transmission to the recipient Diode, thereby producing a chromatographic peak for each specific wavelength.
Waters Acquity UPLC H-Class FLR
Figure 2.2.1: Instrument diagram for the H-Class and I-Class systems. Buffer solutions (up to 4) are steadily pumped through the instrument, consisting of a 6-port valve system, adsorbent column, detectors, and terminates at fraction collection or waste. Samples within the autosampler are loaded into the sample loop via positive displacement from the syringe. The rotation of the valve exposes the sample loop, thereby carrying the sample downstream to the column. Within the column, molecular constituents are separated by adsorption -- a function of the buffers, column, sample properties and temperature. Analytes are continuously carried downstream to the detectors, where they can elicit quantifiable peak signals.
Figure 2.2.2: Principle of Ultraviolet absorbance detection. Filtered light (with a set wavelength) passes through the flow downstream of the column. Presence of light-absorbing molecules will result in less light transmission to the recipient Diode, thereby producing a chromatographic peak.
Figure 2.2.3: Principle of Fluorescence detection. Filtered light (with a set wavelength) is used to excite the flow downstream of the column. The resulting light is filtered and measured with a photo-multiplier tube.
Waters Acquity UPLC H-Class ELS
Figure 2.3.1: Instrument diagram for the H-Class and I-Class systems. Buffer solutions (up to 4) are steadily pumped through the instrument, consisting of a 6-port valve system, adsorbent column, detectors, and terminates at fraction collection or waste. Samples within the autosampler are loaded into the sample loop via positive displacement from the syringe. The rotation of the valve exposes the sample loop, thereby carrying the sample downstream to the column. Within the column, molecular constituents are separated by adsorption -- a function of the buffers, column, sample properties and temperature. Analytes are continuously carried downstream to the detectors, where they can elicit quantifiable peak signals.
Figure 2.3.2: Principle of Ultraviolet absorbance detection. Filtered light (with a set wavelength) passes through the flow downstream of the column. Presence of light-absorbing molecules will result in less light transmission to the recipient Diode, thereby producing a chromatographic peak.
Figure 2.3.3: Principle of Evaporative Light Scattering detection. The column effluent is nebulized -- formed into droplets through flowing nitrogen sheath gas and surrounding heat. Incident light is scattered based on the size of the droplets, where the size is a function of analyte concentration within. The resulting scattering is measured with a photo-multiplier tube.
Waters Acquity UPLC I-Class PDA
Figure 2.4.1: Instrument diagram for the H-Class and I-Class systems. Buffer solutions (up to 4) are steadily pumped through the instrument, consisting of a 6-port valve system, adsorbent column, detectors, and terminates at fraction collection or waste. Samples within the autosampler are loaded into the sample loop via positive displacement from the syringe. The rotation of the valve exposes the sample loop, thereby carrying the sample downstream to the column. Within the column, molecular constituents are separated by adsorption -- a function of the buffers, column, sample properties and temperature. Analytes are continuously carried downstream to the detectors, where they can elicit quantifiable peak signals.
Figure 2.4.2: Principle of PhotodiodeArray detection. Filtered light (across a range of wavelengths) passes through the flow downstream of the column. Presence of light-absorbing molecules will result in less light transmission to the recipient Diode, thereby producing a chromatographic peak for each specific wavelength.
UltiMate 3000
Figure 2.5.1: Instrument diagram for the Ultimate 3000 system. Buffer solutions are steadily pumped through the instrument, consisting of a 6-port valve system, adsorbent column, detectors, and terminates at fraction collection or waste. Samples within the autosampler are loaded into the sample loop via positive displacement from the syringe. The rotation of the valve exposes the sample loop, thereby carrying the sample downstream to the column. Within the column, molecular constituents are separated by adsorption -- a function of the buffers, column, sample properties and temperature. Analytes are continuously carried downstream to the detectors, where they can elicit quantifiable peak signals. Separated analytes can be collected based on the properties of these resulting peaks.
Figure 2.5.2: Principle of Ultraviolet absorbance detection. Filtered light (with a set wavelength) passes through the flow downstream of the column. Presence of light-absorbing molecules will result in less light transmission to the recipient Diode, thereby producing a chromatographic peak.
UltiMate 3000 with FLR Detector
Figure 2.6.1: Instrument diagram for the Ultimate 3000 system. Buffer solutions are steadily pumped through the instrument, consisting of a 6-port valve system, adsorbent column, detectors, and terminates at fraction collection or waste. Samples within the autosampler are loaded into the sample loop via positive displacement from the syringe. The rotation of the valve exposes the sample loop, thereby carrying the sample downstream to the column. Within the column, molecular constituents are separated by adsorption -- a function of the buffers, column, sample properties and temperature. Analytes are continuously carried downstream to the detectors, where they can elicit quantifiable peak signals. Separated analytes can be collected based on the properties of these resulting peaks.
Figure 2.6.2: Principle of Ultraviolet absorbance detection. Filtered light (with a set wavelength) passes through the flow downstream of the column. Presence of light-absorbing molecules will result in less light transmission to the recipient Diode, thereby producing a chromatographic peak.
Figure 2.6.3: Principle of Fluorescence detection. The light beam from the Xenon lamp focused by the lens transmits through the excitation monochromator. The light of the user-selected wavelength is used to excite the flow downstream of the column. The excitation light stimulates the sample to emit fluorescence light. After it is focused by the lens, the emission light is filtered with an emission monochromator to select the light of the user-selected wavelength. A cut-off filter, which lets only light above a certain wavelength pass, is located before the emission monochromator to achieve even better sensitivity. Finally, the filtered emission light is measured with a double-photomultiplier tube (PMT) system to cover the full spectra range from ultraviolet to near-infrared region (220 – 900 nm).
UltiMate 3000 with MALS-DLS-RI Detector
Figure 2.7.1: Instrument diagram for the Ultimate 3000 system. Buffer solutions are steadily pumped through the instrument, consisting of a 6-port valve system, adsorbent column, detectors, and terminates at fraction collection or waste. Samples within the autosampler are loaded into the sample loop via positive displacement from the syringe. The rotation of the valve exposes the sample loop, thereby carrying the sample downstream to the column. Within the column, molecular constituents are separated by adsorption -- a function of the buffers, column, sample properties and temperature. Analytes are continuously carried downstream to the detectors, where they can elicit quantifiable peak signals. Separated analytes can be collected based on the properties of these resulting peaks.
Figure 2.7.2: Principle of Ultraviolet absorbance detection. Filtered light (with a set wavelength) passes through the flow downstream of the column. Presence of light-absorbing molecules will result in less light transmission to the recipient Diode, thereby producing a chromatographic peak.
Figure 2.7.3: Principle of Multi-Angle static Light Scattering (MALS) and Dynamic Light Scattering (DLS) detection. The flow downstream of the column passes through the MALS detector, which is equipped with the DLS detection module. 658 nm laser light is scattered by the nanoparticles or biomacromolecules in the sample inside the flow cell. The intensities of the scattered light beams at different directions are measured by 17 MALS photodiode detectors. Meanwhile, the fluctuation of the scattered light beam is measured by 1 DLS photodiode detector. The DLS photodiode detector has smaller surface area than the MALS photodiode detector to allow accurate measurement of light intensity fluctuation. With the concentration information of the sample, the results from MALS and DLS detection can provide analysis of molar mass, radius of gyration and hydrodynamic radius for copolymers and protein conjugates (glycoproteins, PEGylated proteins, surfactant-bound membrane proteins, etc).
Figure 2.7.4: Principle of refractive index (RI) detection.There are two flow cells inside the RI detector: sample flow cell and reference flow cell.When the reference loading valves are open, the flow downstream of the column is loaded into the two flow cells simultaneously. When the valves are closed,only the sample flow cell is loaded while the reference flow remains stable.The light source of 600 nm passes through the sample flow cell and then the reference flow cell and the refracted light is measured by a photodiode.Under RefractiveIndex mode,the valves are kept open and both flow cells are loaded with the sample.The displacement of the light beam is measured and used to determine the absolute refractive index of the sample. Under DifferentialRefractiveIndex mode,the reference cell is loaded with the desired solvent at the beginning of the experiment and then the valves are closed.The shift of light beam direction is measured and used to determine the differential refractive index of the sample and the solvent.
UltiMate 3000 with PCM Detector
Figure 2.8.1: Instrument diagram for the Ultimate 3000 system. Buffer solutions are steadily pumped through the instrument, consisting of a 6-port valve system, adsorbent column, detectors, and terminates at fraction collection or waste. Samples within the autosampler are loaded into the sample loop via positive displacement from the syringe. The rotation of the valve exposes the sample loop, thereby carrying the sample downstream to the column. Within the column, molecular constituents are separated by adsorption -- a function of the buffers, column, sample properties and temperature. Analytes are continuously carried downstream to the detectors, where they can elicit quantifiable peak signals. Separated analytes can be collected based on the properties of these resulting peaks.
Figure 2.8.2: Principle of Ultraviolet absorbance detection. Filtered light (with a set wavelength) passes through the flow downstream of the column. Presence of light-absorbing molecules will result in less light transmission to the recipient Diode, thereby producing a chromatographic peak.
Figure 2.8.3: Principle of pH and conductivity detection. The flow downstream of the column passes through the conductivity flow cell and the pH flow cell sequentially. Conductivity is monitored with the resulting amperometric signals while alternating voltage is applied on the conductivity probe. pH is measured by the generated voltage between the working electrode and the reference electrode, which is directly proportional to the pH of the solution in the pH flow cell. Temperature sensors are presented in both flow cells to achieve automatic temperature compensation in the pH and conductivity calculation.
Agilent 1290 Infinity II LC System
Figure 2.9.1: Instrument diagram for the Agilent 1290 Infinity system. Buffer solutions are steadily pumped through the instrument, consisting of a 6-port valve system, adsorbent column, detectors, and terminates at fraction collection or waste. Samples within the autosampler are loaded into the sample loop via positive displacement from the syringe. The rotation of the valve exposes the sample loop, thereby carrying the sample downstream to the column. Within the column, molecular constituents are separated by adsorption -- a function of the buffers, column, sample properties and temperature. Analytes are continuously carried downstream to the detectors, where they can elicit quantifiable peak signals. Separated analytes can be collected based on the properties of these resulting peaks.
Figure 2.9.2: Agilent 1290 Infinity plumbing diagram. The solvent flow from each pump head is merged at a t-piece in pairs. Solvents A/C share the same flow path from here on as well as B/D. Joint flow from from each pump pair then travels through a pressure sensor and a purge valve before it is combined at the manifold where it then moves on to the mixer and an autosampler. This configuration does not allow for having a gradient of A/C or B/D and only mixes between the two sides are possible.
Figure 2.9.3: Principle of PhotodiodeArray detection. Filtered light (across a range of wavelengths) passes through the flow downstream of the column. Presence of light-absorbing molecules will result in less light transmission to the recipient Diode, thereby producing a chromatographic peak for each specific wavelength.
Agilent 1260 Infinity II Semi-Preparative HPLC with UV/Vis Diode Array Detector
Figure 2.10.1: Instrument diagram for the Agilent 1260 Infinity II Semi-Preparative HPLC system which outlines the flow of buffer solutions through the instrument including a 6-port valve, an adsorbent column, and detectors, ultimately leading to either fraction collection or waste. Buffer solutions are continuously pumped through the instrument. Samples in the autosampler are introduced into the sample loop via positive displacement from the syringe. When the valve rotates, it exposes the sample loop, allowing the sample to be carried downstream to the column by the buffer flow. Inside the column, molecular constituents are separated by adsorption, influenced by the buffers, the column, the sample properties, and the temperature. Analytes are then carried downstream to the detectors, where they produce quantifiable peak signals. These separated analytes can be collected based on the characteristics of the resulting peaks.
Figure 2.10.2: Agilent 1260 Infinity II Semi-Preparative HPLC system quaternary pump plumbing diagram. The flow of each buffer is controlled independently with four separate pump heads. Each flow passes through a pressure sensor and a purge valve before combined at the manifold. The combined flow then moves on to the mixer and subsequently to the autosampler of the instrument.
Figure 2.10.3: Principle of PhotodiodeArray detection. Filtered light (across a range of wavelengths) passes through the flow downstream of the column. Presence of light-absorbing molecules will result in less light transmission to the recipient Diode, thereby producing a chromatographic peak for each specific wavelength.
Agilent 1260 Infinity II Semi-Preparative HPLC with UV/Vis Diode Array and Fluorescence Detectors
Figure 2.11.1: Instrument diagram for the Agilent 1260 Infinity II Semi-Preparative HPLC system which outlines the flow of buffer solutions through the instrument including a 6-port valve, an adsorbent column, and detectors, ultimately leading to either fraction collection or waste. Buffer solutions are continuously pumped through the instrument. Samples in the autosampler are introduced into the sample loop via positive displacement from the syringe. When the valve rotates, it exposes the sample loop, allowing the sample to be carried downstream to the column by the buffer flow. Inside the column, molecular constituents are separated by adsorption, influenced by the buffers, the column, the sample properties, and the temperature. Analytes are then carried downstream to the detectors, where they produce quantifiable peak signals. These separated analytes can be collected based on the characteristics of the resulting peaks.
Figure 2.11.2: Agilent 1260 Infinity II Semi-Preparative HPLC system quaternary pump plumbing diagram. The flow of each buffer is controlled independently with four separate pump heads. Each flow passes through a pressure sensor and a purge valve before combined at the manifold. The combined flow then moves on to the mixer and subsequently to the autosampler of the instrument.
Figure 2.11.3: Principle of PhotodiodeArray detection. Filtered light (across a range of wavelengths) passes through the flow downstream of the column. Presence of light-absorbing molecules will result in less light transmission to the recipient Diode, thereby producing a chromatographic peak for each specific wavelength.
Figure 2.11.4: Principle of Fluorescence detection. The light beam from the Xenon lamp focused by the lens transmits through the excitation monochromator (200 – 1200 nm with 20 nm fixed bandwidth). The light of the user-selected wavelength is used to excite the flow downstream of the column. The excitation light stimulates the sample to emit fluorescence light. After it is focused by the lens, the emission light is filtered with an emission monochromator (200 – 1200 nm with 20 nm fixed bandwidth) to select the light of the user-selected wavelength. Finally, the filtered emission light is measured with a double-photomultiplier tube (PMT) system to cover the full spectra range from ultraviolet to near-infrared region (200 – 1200 nm).
Agilent 1260 Infinity II Semi-Preparative HPLC with MALS-DLS-RI Detector
Figure 2.12.1: Instrument diagram for the Agilent 1260 Infinity II Semi-Preparative HPLC system which outlines the flow of buffer solutions through the instrument including a 6-port valve, an adsorbent column, and detectors, ultimately leading to either fraction collection or waste. Buffer solutions are continuously pumped through the instrument. Samples in the autosampler are introduced into the sample loop via positive displacement from the syringe. When the valve rotates, it exposes the sample loop, allowing the sample to be carried downstream to the column by the buffer flow. Inside the column, molecular constituents are separated by adsorption, influenced by the buffers, the column, the sample properties, and the temperature. Analytes are then carried downstream to the detectors, where they produce quantifiable peak signals. These separated analytes can be collected based on the characteristics of the resulting peaks.
Figure 2.12.2: Agilent 1260 Infinity II Semi-Preparative HPLC system quaternary pump plumbing diagram. The flow of each buffer is controlled independently with four separate pump heads. Each flow passes through a pressure sensor and a purge valve before combined at the manifold. The combined flow then moves on to the mixer and subsequently to the autosampler of the instrument.
Figure 2.12.3: Principle of PhotodiodeArray detection. Filtered light (across a range of wavelengths) passes through the flow downstream of the column. Presence of light-absorbing molecules will result in less light transmission to the recipient Diode, thereby producing a chromatographic peak for each specific wavelength.
Figure 2.12.4: Principle of Multi-Angle static Light Scattering (MALS) and Dynamic Light Scattering (DLS) detection. The flow downstream of the column passes through the MALS detector, which is equipped with the DLS detection module. 658 nm laser light is scattered by the nanoparticles or biomacromolecules in the sample inside the flow cell. The intensities of the scattered light beams at different directions are measured by 17 MALS photodiode detectors. Meanwhile, the fluctuation of the scattered light beam is measured by 1 DLS photodiode detector. The DLS photodiode detector has smaller surface area than the MALS photodiode detector to allow accurate measurement of light intensity fluctuation. With the concentration information of the sample, the results from MALS and DLS detection can provide analysis of molar mass, radius of gyration and hydrodynamic radius for copolymers and protein conjugates (glycoproteins, PEGylated proteins, surfactant-bound membrane proteins, etc).
Figure 2.12.5: Principle of refractive index (RI) detection.There are two flow cells inside the RI detector: sample flow cell and reference flow cell.When the reference loading valves are open, the flow downstream of the column is loaded into the two flow cells simultaneously. When the valves are closed,only the sample flow cell is loaded while the reference flow remains stable.The light source of 600 nm passes through the sample flow cell and then the reference flow cell and the refracted light is measured by a photodiode.Under RefractiveIndex mode,the valves are kept open and both flow cells are loaded with the sample.The displacement of the light beam is measured and used to determine the absolute refractive index of the sample. Under DifferentialRefractiveIndex mode,the reference cell is loaded with the desired solvent at the beginning of the experiment and then the valves are closed.The shift of light beam direction is measured and used to determine the differential refractive index of the sample and the solvent.
Experiment Options
General
Instrument
A list of one or more measurement and collection devices to run the experiment on and that satisfy the Scale and Detector options.
Default Calculation: For FractionCollection option specification and/or for Scale->Preparative or SemiPreparative, automatically set to instrument models that have fraction collection capabilities. Otherwise, automatically set to instrument models that meet the requested Detector.
Pattern Description: An object of type or subtype Model[Instrument, HPLC] or Object[Instrument, HPLC] or list of one or more an object of type or subtype Model[Instrument, HPLC] entries or Null.
Programmatic Pattern: ((ObjectP[{Model[Instrument, HPLC], Object[Instrument, HPLC]}] | {ObjectP[Model[Instrument, HPLC]]..}) | Automatic) | Null
Scale
The output of the experiment. Preparative and SemiPreparative indicates that effluent is to be collected by fractions. Analytical indicates that specific measurements will be employed and new SamplesOut will not be generated (e.g the absorbance of the flow with injected sample for a given wavelength).
Default Calculation: If any fraction collection options are specified and injection volume is greater that 500 uL, then Scale -> Preparative; if fraction collection options are specified and injection volume is less than or equal to 500uL, then Scale -> SemiPreparative; otherwise, Scale -> Analytical.
SeparationMode
The category of method used to elicit differential column retention due to interaction between molecules in the mobile phase with those on the stationary phase (column).
Default Calculation: Automatically set to match the Separation Mode listed with the provided column.
Detector
The type of measurement to employ. Options include Pressure (measures the pump pressure) , Temperature (measures the temperature of the column oven), UVVis (measures the absorbance of a single wavelength of light), PhotoDiodeArray (measures the absorbance of a range of wavelengths), Fluorescence (measures the emitted light from samples after light excitation),pH, Conductance, MultiAngleLightScattering (measures the scattered light intensity at different angles), DynamicLightScattering (measures the scattered light fluctuation), RefractiveIndex (measures how fast light travels through the sample) and EvaporativeLightScattering (separates the flow into airborne droplets and measures the light scattering).
Default Calculation: Automatically set to the detector(s) available for the first selected instrument. For example, if Agilent 1290 Infinity II Instrument is requested, the Detector option will include Pressure and PhotoDiodeArray.
Pattern Description: A selection of one or more of Pressure, Temperature, Conductance, Fluorescence, EvaporativeLightScattering, UVVis, PhotoDiodeArray, CircularDichroism, RefractiveIndex, pH, MultiAngleLightScattering, or DynamicLightScattering.
Programmatic Pattern: DuplicateFreeListableP[Pressure | Temperature | Conductance | Fluorescence | EvaporativeLightScattering | UVVis | PhotoDiodeArray | CircularDichroism | RefractiveIndex | pH | MultiAngleLightScattering | DynamicLightScattering] | Automatic
ColumnSelection
Indicates if multiple different columns will be employed for different samples during the run. All columns are installed during the beginning of the run and the valve on the instrument allows to switch between them automatically.
Default Calculation: If ColumnSelector and InjectionTable are not specified, automatically set to False. If InjectionTable is set but with only one set of column(s), automatically set to False. Otherwise, set to True.
ColumnPosition
The position of the column selector valve and the desired column configuration that will be used for each sample as it is injected.
Default Calculation: If InjectionTable is specified, automatically set from the Column Position entry for the sample. Otherwise set to PositionA.
Pattern Description: PositionA, PositionB, PositionC, PositionD, PositionE, PositionF, PositionG, or PositionH.
GuardColumn
The protective device placed in the flow path before the Column in order to adsorb fouling contaminants and, thus, preserve the Column lifetime.
Default Calculation: Automatically set from the column model's PreferredGuardColumn. If Column is Null, GuardColumn is automatically set to Null.
Pattern Description: An object of type or subtype Model[Item, Column], Object[Item, Column], Model[Item, Cartridge, Column], or Object[Item, Cartridge, Column] or Null.
Programmatic Pattern: (ObjectP[{Model[Item, Column], Object[Item, Column], Model[Item, Cartridge, Column], Object[Item, Cartridge, Column]}] | Automatic) | Null
GuardColumnOrientation
The position of the GuardColumn with respect to the Column, SecondaryColumn and TertiaryColumn. Forward indicates that the GuardColumn will be placed in front of the Column, SecondaryColumn and TertiaryColumn. If a Column is specified and GuardColumnOrientation is Reverse, the GuardColumn will be placed after the Column, SecondaryColumn, and/or TertiaryColumn in the flow path which is typically performed during column cleaning.
Column
The item containing the stationary phase through which the mobile phase and input samples flow. It adsorbs and separates the molecules within the sample based on the properties of the mobile phase, Samples, Column material, and the desired column temperature in the specified InjectionTable.
Default Calculation: Automatically set to a column model compatible for the instrument selected and specified separation Mode.
Pattern Description: An object of type or subtype Model[Item, Column] or Object[Item, Column] or Null.
ColumnOrientation
The direction of the Column with respect to the flow. Forward indicates that the Column will be placed in the direction indicated by the column manufacturer for standard operation. Reverse indicates that the Column will be placed in the opposite direction indicated by the column manufacturer for standard operation. This also specifies the orientation of secondary and tertiary columns if provided.
SecondaryColumn
The additional stationary phase through which the mobile phase and input samples flow. The SecondaryColumn selectively adsorb analytes and is connected to flow path, downstream of the Column.
Pattern Description: An object of type or subtype Model[Item, Column] or Object[Item, Column] or Null.
TertiaryColumn
The additional stationary phase through which the mobile phase and input samples flow. The TertiaryColumn selectively adsorb analytes and is connected to flow path, downstream of the SecondaryColumn.
Pattern Description: An object of type or subtype Model[Item, Column] or Object[Item, Column] or Null.
IncubateColumn
Indicates if the columns are placed inside the column oven compartment of the HPLC instrument during the experiment. If set to False, the columns are placed on a rack outside the column oven under ambient temperature.
Default Calculation: Automatically set to False if the selected connection of GuardColumn, Column, SecondaryColumn, and TertiaryColumn cannot fit into the column oven compartment of the Instrument. Otherwise set to True.
BufferA
A solvent or buffer placed in the 'A' bottle as shown in Figure 2.1.1 - 2.9.1 of ExperimentHPLC help file, pumped through the instrument as part of the mobile phase, the compositions of which is determined by the GradientA option.
Default Calculation: Automatically set from the SeparationMode (e.g. Water buffer if ReversePhase) option or the objects specified in the Gradient option.
Pattern Description: An object of type or subtype Object[Sample] or Model[Sample] or a prepared sample.
BufferB
A solvent or buffer placed in the 'B' bottle as shown in Figure 2.1.1 - 2.9.1 of ExperimentHPLC help file, pumped through the instrument as part of the mobile phase, the compositions of which is determined by the GradientB option.
Default Calculation: Automatically set from SeparationMode (e.g. Acetonitrile buffer if ReversePhase) or the objects specified by the Gradient option.
Pattern Description: An object of type or subtype Object[Sample] or Model[Sample] or a prepared sample.
BufferC
A solvent or buffer placed in the 'C' bottle as shown in Figure 2.1.1 - 2.9.1 of ExperimentHPLC help file, pumped through the instrument as part of the mobile phase, the compositions of which is determined by the GradientC option.
Default Calculation: Automatically set from the SeparationMode option or the objects specified in the Gradient option.
Pattern Description: An object of type or subtype Object[Sample] or Model[Sample] or a prepared sample or Null.
BufferD
A solvent or buffer placed in the 'D' bottle as shown in Figure 2.1.1 - 2.5.1 of ExperimentHPLC help file, pumped through the instrument as part of the mobile phase, the compositions of which is determined by the GradientD option.
Default Calculation: If the specified Instrument's pump does not support Buffer D, automatically set to Null. Otherwise, set from the SeparationMode option or the objects specified in the Gradient option.
Pattern Description: An object of type or subtype Object[Sample] or Model[Sample] or a prepared sample or Null.
Column Installation
ColumnSelector
The set of all the columns loaded into the Instrument's column selector and referenced in Column, SecondaryColumn, TertiaryColumn. The Serial configuration indicates one fluid line for all the samples, Standard and Blank. The Selector configuration indicates use of a column selector where the column line is programmatically switchable between samples.
Default Calculation: If ColumnSelection is False, set to match the values in Column, SecondaryColumn, TertiaryColumn, ColumnOrientation, GuardColumn and GuardColumnOrientation options.
Pattern Description: {Column Position, Guard Column, Guard Column Orientation, Column, Column Orientation, Secondary Column, Tertiary Column}
Programmatic Pattern: {ColumnPositionP | Automatic | Null, ObjectP[{Model[Item, Column], Object[Item, Column]}] | (Automatic | Null), ColumnOrientationP | (Automatic | Null), ObjectP[{Model[Item, Column], Object[Item, Column]}] | (Automatic | Null), ColumnOrientationP | (Automatic | Null), ObjectP[{Model[Item, Column], Object[Item, Column]}] | (Automatic | Null), ObjectP[{Model[Item, Column], Object[Item, Column]}] | (Automatic | Null)} | Automatic
Separation
ColumnStorageBuffer
The solvent or gradient at the end of the column flush in which the column will be stored in long term after removed from the instrument. The provided solvent option must match one of the buffers used in the experiment and the column flush will end with 100% gradient of the selected buffer. If a gradient with percents of buffers is specified, the column flush will end with the specified gradient composition.
Default Calculation: Automatically set to the last gradient composition of the ColumnFlushGradient option if provided. Otherwise set from the StorageBuffer field from the Model[Item,Column] specified for the specified Column, if it is one of the buffers specified in the protocol.
Programmatic Pattern: (((ObjectP[{Object[Sample], Model[Sample]}] | _String) | {RangeP[0*Percent, 100*Percent], RangeP[0*Percent, 100*Percent], RangeP[0*Percent, 100*Percent], RangeP[0*Percent, 100*Percent]}) | Automatic) | Null
GradientA
The composition of BufferA within the flow, defined for specific time points. The composition is linearly interpolated for the intervening periods between the defined time points. For example for GradientA->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferA in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If Gradient option is specified, set from it or implicitly determined from the GradientB, GradientC, and GradientD options such that the total amounts to 100%.
Programmatic Pattern: (RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic
GradientB
The composition of BufferB within the flow, defined for specific time points. The composition is linearly interpolated for the intervening periods between the defined time points. For example for GradientB->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferB in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If Gradient option is specified, set from it or implicitly determined from the GradientA, GradientC, and GradientD options such that the total amounts to 100%.
Programmatic Pattern: (RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic
GradientC
The composition of BufferC within the flow, defined for specific time points. The composition is linearly interpolated for the intervening periods between the defined time points. For example for GradientC->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferC in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If Gradient option is specified, set from it or implicitly determined from the GradientA, GradientB, and GradientD options such that the total amounts to 100%.
Programmatic Pattern: (RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic
GradientD
The composition of BufferD within the flow, defined for specific time points. The composition is linearly interpolated for the intervening periods between the defined time points. For example for GradientD->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferD in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If Gradient option is specified, set from it or implicitly determined from the GradientA, GradientB, and GradientC options such that the total amounts to 100%.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic) | Null
FlowRate
The net speed of the fluid flowing through the pump inclusive of the composition of BufferA, BufferB, BufferC, and BufferD specified in the gradient options. This speed is linearly interpolated such that consecutive entries of {Time, Flow Rate} will define the intervening fluid speed. For example, {{0 Minute, 0.3 Milliliter/Minute},{30 Minute, 0.5 Milliliter/Minute}} means flow rate of 0.4 Milliliter/Minute at 15 minutes into the run.
Default Calculation: If Gradient option is specified, automatically set from the method given in the Gradient option. If NominalFlowRate of the column model is specified, set to lesser of the NominalFlowRate for each of the columns, guard columns or the instrument's MaxFlowRate. Otherwise set to 1 Milliliter / Minute.
Pattern Description: Greater than or equal to 0 milliliters per minute and less than or equal to 200 milliliters per minute or list of one or more {Time, Flow Rate} entries.
Programmatic Pattern: (RangeP[0*(Milliliter/Minute), 200*(Milliliter/Minute)] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*(Milliliter/Minute), 200*(Milliliter/Minute)]}..}) | Automatic
MaxAcceleration
When ramping up the FlowRate of solvent through the instrument, the maximum allowed change per time in the FlowRate.
Default Calculation: For Waters and Agilent instruments, automatically set to the lowest value from Max the Column, Instrument, and GuardColumn models. For other instruments, automatically set to Null.
Gradient
The composition of different specified buffers in BufferA, BufferB, BufferC and BufferD over time in the fluid flow. Specific parameters of a gradient object can be overridden by specific options. Differential Refractive Index Reference Loading refers to the HPLC refractive index loading reference values as shown in the Figure 2.7.4. When open, the flow downstream of the column is loaded into the two flow cells simultaneously.
Default Calculation: Automatically set to best meet all the Gradient options (e.g. GradientA, GradientB, GradientC, GradientD, FlowRate).
Pattern Description: An object of type or subtype Object[Method, Gradient] or list of one or more {Time, Buffer A Composition, Buffer B Composition, Buffer C Composition, Buffer D Composition, Flow Rate, Differential Refractive Index Reference Loading} entries.
Programmatic Pattern: (ObjectP[Object[Method, Gradient]] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent], RangeP[0*Percent, 100*Percent], RangeP[0*Percent, 100*Percent], RangeP[0*Percent, 100*Percent], RangeP[0*(Milliliter/Minute), 200*(Milliliter/Minute)], Open | Closed | None | Automatic}..}) | Automatic
Sample Parameters
NumberOfReplicates
The number of times to repeat measurements on each provided sample(s). If Aliquot -> True, this also indicates the number of times each provided sample will be aliquoted. For experiment samples {A,B,C} if NumberOfReplicates is specified as 3, the order of samples to run on the instrument will be {A,A,A,B,B,B,C,C,C}.
Pattern Description: Greater than or equal to 1 and less than or equal to 96 in increments of 1 or Null.
InjectionTable
The order of Sample, Standard and Blank sample injected into the Instrument during measurement and/or collection. This also includes the priming and flushing of the column(s).
Default Calculation: Samples are inserted in the order of the input samples based with the number of replicates. Standard and Blank samples are inserted based on the determination of StandardFrequency and BlankFrequency options. For example, StandardFrequency -> FirstAndLast and BlankFrequency -> Null result in Standard samples injected first, then samples, and then the Standard sample set again at the end. Column priming is inserted at the beginning and repeated according to ColumnPrimeFrequency. Column flushing is inserted at the end.
Pattern Description: List of one or more {Type, Sample, InjectionVolume, Column Position, Column Temperature, Gradient} entries.
Programmatic Pattern: {{Standard | Sample | Blank | ColumnPrime | ColumnFlush, (ObjectP[{Model[Sample], Object[Sample]}] | _String) | (Automatic | Null), RangeP[0*Microliter, 16*Milliliter] | (Automatic | Null), ColumnPositionP | Automatic, RangeP[5*Celsius, 90*Celsius] | (Ambient | Automatic), ObjectP[Object[Method, Gradient]] | Automatic}..} | Automatic
SampleTemperature
The temperature of the chamber in which the samples, Standard, and Blank are stored while waiting for the Injection.
Pattern Description: Ambient or greater than or equal to 5 degrees Celsius and less than or equal to 40 degrees Celsius.
ColumnTemperature
Default Calculation: Automatically set to the corresponding gradient temperature specified in the Gradient option or the column temperature for the sample in the InjectionTable option; otherwise, set to Ambient (no column oven temperature control).
Pattern Description: Ambient or greater than or equal to 5 degrees Celsius and less than or equal to 90 degrees Celsius.
InjectionVolume
Default Calculation: Automatically defaults to the lesser of 10 uL or 90% of the available sample volume for Analytical measurement, lesser of 500 uL or 90% of the available sample volume for Semipreparative measurement and lesser of 5mL or 90% of available sample volume for Preparative measurement.
Pattern Description: Greater than or equal to 0 microliters and less than or equal to 16 milliliters.
NeedleWashSolution
The solvent used to wash the injection needle before each sample introduction. For Dionex instruments, this is the same as BufferC and cannot be defined separately.
Default Calculation: If the instrument shares NeedleWashSolution with BufferC, automatically set to specified BufferC. Otherwise, defaults to Model[Sample, "Milli-Q water"] for IonExchange and SizeExclusion SeparationType or Model[Sample, StockSolution, "20% Methanol in MilliQ Water"] for other SeparationType.
Pattern Description: An object of type or subtype Object[Sample] or Model[Sample] or a prepared sample.
Detection
AbsorbanceWavelength
The wavelength of light passed through the flow cell for the UVVis Detector. For PhotoDiodeArray Detector, a 3D data is generated from a spectrum of light passing through the flow cell. Absorbance wavelength in that case represents the wavelength at which a 2D data slice is generated from the 3D data.
Default Calculation: If a UVVis Detector is selected or available on the selected instrument, automatically set to the absorbance wavelength corresponding to the maximum extinction coefficient from the ExtinctionCoefficients field in the identity model of the samples specified. If no ExtinctionCoefficients available, automatically set to to 260 Nanometer for oligomers or 280 Nanometer for proteins. If a PhotoDiodeArray Detector is selected or available on the selected Instrument, automatically set to All.
Programmatic Pattern: ((RangeP[190*Nanometer, 950*Nanometer] | All | RangeP[190*Nanometer, 950*Nanometer] ;; RangeP[200*Nanometer, 950*Nanometer]) | Automatic) | Null
WavelengthResolution
The increment in wavelength for the range of wavelength of light passed through the flow for absorbance measurement of PhotoDiodeArray Detector.
Default Calculation: If a PhotoDiodeArray Detector is selected or available on the selected Instrument, automatically set to 2.4 Nanometer.
Pattern Description: Greater than or equal to 0.1 nanometers and less than or equal to 12. nanometers or Null.
UVFilter
Indicates if UV wavelengths (less than 210 nm) should be blocked from being transmitted through the sample for the PhotoDiodeArray Detector.
Default Calculation: If a PhotoDiodeArray Detector is selected or available on the selected Instrument, automatically set to False.
AbsorbanceSamplingRate
The number of times an absorbance measurement is made per second by the detector on the selected instrument. Lower values will be less susceptible to noise but will record less frequently across time.
Default Calculation: If a UVVis Detector or PhotoDiodeArray Detector is selected or available on the selected instrument, automatically set to 20/Second .
Pattern Description: Greater than or equal to 1 reciprocal second and less than or equal to 120 reciprocal seconds or Null.
ExcitationWavelength
The wavelength(s) of light that is used to excite fluorescence in the samples when passed through the Fluorescence Detector.
Default Calculation: If Fluorescence Detector is selected, automatically set from the FluorescenceExcitationMaximums field in the identity Model of the sample specified.
Programmatic Pattern: ((RangeP[200*Nanometer, 1200*Nanometer] | {RangeP[200*Nanometer, 890*Nanometer]..}) | Automatic) | Null
EmissionWavelength
The wavelength(s) of light at which fluorescence emitted from the sample is measured in the Fluorescence Detector.
Default Calculation: If Fluorescence Detector is selected, automatically set from the FluorescenceEmissionMaximums field in the identity Model of the sample specified.
Programmatic Pattern: ((RangeP[200*Nanometer, 1200*Nanometer] | {RangeP[200*Nanometer, 890*Nanometer]..}) | Automatic) | Null
EmissionCutOffFilter
The cut-off wavelength to pre-select the emitted light from the sample and allow only the light with wavelength above the desired value to pass, before the light enters emission monochromator for final wavelength selection for Ultimate 3000 with FLR Detector. If set to None, no cut-off filter is used.
Default Calculation: If a Fluorescence Detector with a cut-off filter wheel is selected, automatically set to None.
Pattern Description: 280 nanometers, 370 nanometers, 435 nanometers, 530 nanometers, or None or Null.
FluorescenceGain
For each ExcitationWavelength/EmissionWavelength pair, the signal amplification factor which modulates the percentage of maximum voltage that can be applied to the Photomultiplier Tube of the Fluorescence Detector. Linear increase in voltage applied to the Photomultiplier tube leads to an exponential change in RFU signal. Variable Fluorescence Sensitivity implies a different fluorescence sensitivity for each Excitation/Emission Wavelength pair.
Default Calculation: If the "Ultimate 3000 with FLR Detector" or "Waters Acquity UPLC H-Class FLR" instrument is selected, automatically set to 100 Percent. If the "Agilent 1260 Infinity II Semi-Preparative HPLC with UV/Vis Diode Array and Fluorescence Detectors" instrument is selected, automatically set to 60 Percent.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {RangeP[0*Percent, 100*Percent]..}) | Automatic) | Null
FluorescenceFlowCellTemperature
The temperature that the thermostat inside the fluorescence flow cell of the Fluorescence Detector is set to during the fluorescence measurement of the sample.
Default Calculation: If Fluorescence Detector is selected and temperature control is available on that unit, automatically set to Ambient.
Pattern Description: Ambient or greater than or equal to 25 degrees Celsius and less than or equal to 50 degrees Celsius or Null.
LightScatteringLaserPower
The laser power filter used in the Multi-Angle static Light Scattering (MALS) and Dynamic Light Scattering (DLS) Detector. 100% means that no filter is being used to restrict the laser power.
Default Calculation: If MultiAngleLightScattering Detector and/or DynamicLightScattering Detector are selected, automatically set to 100 Percent.
Pattern Description: Greater than or equal to 10 percent and less than or equal to 100 percent or Null.
LightScatteringFlowCellTemperature
The temperature that the thermostat inside the flow cell of the Detector is set to during the Multi-Angle static Light Scattering (MALS) and/or Dynamic Light Scattering (DLS) measurement of the sample.
Default Calculation: If MultiAngleLightScattering Detector and/or DynamicLightScattering Detector are selected, automatically set to Ambient.
Pattern Description: Ambient or greater than or equal to 20 degrees Celsius and less than or equal to 70 degrees Celsius or Null.
RefractiveIndexMethod
The type of refractive index measurement of the Refractive Index (RI) Detector for the measurement of the sample. When DifferentialRefractiveIndex is selected, the refractive index difference between the flow downstream sample and the reference solvent is measured. See Figure 2.7.4 for more information.
Default Calculation: If RefractiveIndex Detector is selected and Differential Refractive Index Reference Loading is set to Closed in Gradient, automatically set to DifferentialRefractiveIndex. Otherwise automatically set to RefractiveIndex.
RefractiveIndexFlowCellTemperature
The temperature that the thermostat inside the refractive index flow cell of the Refractive Index (RI) Detector is set to during the refractive index measurement of the sample.
Pattern Description: Ambient or greater than or equal to 4 degrees Celsius and less than or equal to 65 degrees Celsius or Null.
pHCalibration
Indicates if 2-point calibration of the pH probe should be performed before the experiment starts. pH And Conductivity calibration is performed monthly every time a qualification procedure is run on the instrument.
LowpHCalibrationBuffer
Default Calculation: Automatically set to Model[Sample, "pH 4.01 Calibration Buffer, Sachets"] if pH Detector is selected and pHCalibration is True.
Pattern Description: An object of type or subtype Object[Sample] or Model[Sample] or a prepared sample or Null.
LowpHCalibrationTarget
The pH of the LowpHCalibrationBuffer that should be used to calibrate the pH probe in the 2-point calibration.
HighpHCalibrationBuffer
Default Calculation: Automatically set to Model[Sample, "pH 7.00 Calibration Buffer, Sachets"] if pH Detector is selected and pHCalibration is True. If HighpHCalibrationTarget is specified, set to a buffer with pH value close to that.
Pattern Description: An object of type or subtype Object[Sample] or Model[Sample] or a prepared sample or Null.
HighpHCalibrationTarget
The pH of the HighpHCalibrationBuffer that should be used to calibrate the pH probe in the 2-point calibration.
pHTemperatureCompensation
Indicates if the measured pH value should be automatically corrected according to the temperature inside the pH flow cell.
ConductivityCalibration
Indicates if 1-point calibration of the conductivity probe should be performed before the experiment starts. pH And Conductivity calibration is performed monthly every time a qualification procedure is run on the instrument.
ConductivityCalibrationBuffer
Default Calculation: Automatically set to Model[Sample, "Conductivity Standard 1413 µS, Sachets"] if Conductivity Detector is selected and ConductivityCalibration is True.
Pattern Description: An object of type or subtype Object[Sample] or Model[Sample] or a prepared sample or Null.
ConductivityCalibrationTarget
The conductivity value of the ConductivityCalibrationBuffer that should be used to calibrate the conductivity probe in the 1-point calibration.
Default Calculation: Automatically set to the Conductivity of the ConductivityCalibrationBuffer's model.
Pattern Description: Greater than or equal to 10 microsiemens per centimeter and less than or equal to 1000 millisiemens per centimeter or Null.
Programmatic Pattern: (RangeP[10*Micro*(Siemens/Centimeter), 1000*Milli*(Siemens/Centimeter)] | Automatic) | Null
ConductivityTemperatureCompensation
Indicates if the measured conductivity value should be automatically corrected according to the temperature inside the conductivity flow cell.
NebulizerGas
Indicates if Nitrogen sheath gas is flowed along with the sample within the EvaporativeLightScattering Detector.
NebulizerGasHeating
Indicates if the sheath gas that carries samples in the EvaporativeLightScattering Detector is heated.
Default Calculation: If EvaporativeLightScattering Detector is selected and NebulizerGas is True, automatically set to True.
NebulizerHeatingPower
The relative magnitude of the heating element used to heat the sheath gas for the EvaporativeLightScattering Detector (Corresponding temperature not measured by the manufacturer). Higher percent values correspond to percent of power going to heating coil.
Default Calculation: If EvaporativeLightScattering Detector is selected and NebulizerGasHeating is True, automatically set to 50 Percent.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or Null.
NebulizerGasPressure
The applied pressure of sheath gas for the EvaporativeLightScattering Detector (Flow rate unmeasured by the manufacturer). Higher pressure (20-60 PSI) corresponds to faster sheath gas flow.
Default Calculation: If EvaporativeLightScattering Detector is selected and NebulizerGas is True, automatically set to 40 PSI.
Pattern Description: Greater than or equal to 20 pounds‐force per inch squared and less than or equal to 60 pounds‐force per inch squared or Null.
DriftTubeTemperature
The set temperature of the chamber thermostat through which the nebulized analytes flow within the EvaporativeLightScattering Detector. The purpose to heat the drift tube is to evaporate any unevaporated solvent remaining in the flow from the nebulizer.
Default Calculation: If EvaporativeLightScattering Detector is selected and NebulizerGas is True, automatically set to 50 Celsius.
Pattern Description: Greater than or equal to 20 degrees Celsius and less than or equal to 100 degrees Celsius or Null.
ELSDGain
The percent of maximum voltage sent to the Photo Multiplier Tube (PMT) for signal amplification for the EvaporativeLightScattering measurement. The percentage value specified here is converted into a unitless factor from 0 to 1000 which the software accepts to modulate the voltage for the PMT.
Default Calculation: If EvaporativeLightScattering Detector is selected and NebulizerGas is True, automatically set to 50 Percent.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or Null.
ELSDSamplingRate
The frequency of evaporative light scattering measurement. Lower values will be less susceptible to noise but will record less frequently across time. Lower or higher values do not affect the y axis of the measurement.
Default Calculation: If EvaporativeLightScattering Detector is selected and NebulizerGas is True, automatically set to 1/Second.
Pattern Description: Greater than or equal to 1 reciprocal second and less than or equal to 80 reciprocal seconds or Null.
Fraction Collection
CollectFractions
Indicates if effluents from the Column should be captured and stored at specific time windows or during detection of peaks (fractions).
Default Calculation: If Scale is Preparative/SemiPreparative or any fraction collection options are specified, set to True. For analytical measurements, set to False.
FractionCollectionDetector
The type of measurement that is used as signal to trigger fraction collection. It corresponds to the type of detector on the instrument.
Default Calculation: If CollectFractions is True, automatically set to the Detector in the Detector option for which the Detector hardware can communicate with the fraction collection (as indicated in the Instrumentation Table of ExperimentHPLC help file).
FractionCollectionContainer
Default Calculation: Automatically set to Model[Container, Plate, "96-well 2mL Deep Well Plate"] for UltiMate 3000 HPLC instruments and to Model[Container, Vessel, "50mL Tube"] for Agilent 1290 Infinity II instrument.
FractionCollectionMethod
The fraction collection method object which describes the conditions for which a fraction is collected. Specific parameters of the object can be overridden by other fraction collection options.
FractionCollectionStartTime
The time at which to start column effluent capture. Time in this case is the duration from the start of sample injection.
Default Calculation: Automatically set from the method specified in the FractionCollectionMethod option, if available. Otherwise set to the second time point of the gradient domains if there are more than two time points, or the first time point if not.
FractionCollectionEndTime
The time to end column effluent capture. Time in this case is the duration from the start of sample injection.
Default Calculation: Automatically inherited from the method specified in the FractionCollectionMethod option. Otherwise set to the last time point of the gradient domains.
FractionCollectionMode
The method by which fractions collection should be triggered (peak detection, a constant threshold, or a fixed fraction time). In Peak detection mode, the fraction collection is triggered when a change in slope of the FractionCollectionDetector signal is observed for a specified PeakSlopeDuration time. In constant Threshold mode, whenever the signal from the FractionCollectionDetector is above the specified value, fraction collection is triggered. In fixed fraction Time mode, fractions are collected during the whole time interval specified.
Default Calculation: Automatically inherited from a method specified by FractionCollectionMethod option, or implicitly resolved from other fraction collection options. If AbsoluteThreshold is specified, set to Threshold. If PeakSlope is specified, set to Peak. If MaxCollectionPeriod is specified, set to Time. Otherwise set to Threshold if CollectFractions is True.
MaxFractionVolume
The maximum amount of sample to be collected in a single fraction. If fraction detection trigger is not off, the collector moves position to the next container. For example, if AbsorbanceThreshold is set to 180 MilliAbsorbanceUnit and at MaxFractionVolume the absorbance value is still above 180 MilliAbsorbanceUnit, the fraction collector continues to collect fractions in the next container in line.
Default Calculation: If FractionCollection is True, automatically set according to the MaxFractionVolume in the method specified by FractionCollectionMethod option, if available. If FractionCollectionContainer is specified, set to MaxVolume of the Model specified. Otherwise, automatically set to 1.8 Milliliter for UltiMate 3000 HPLC instruments and 45 Milliliter for Agilent 1290 Infinity II instrument.
Pattern Description: Greater than or equal to 10 microliters and less than or equal to 50 milliliters or Null.
MaxCollectionPeriod
The amount of time after which a new fraction will be generated (Fraction Collector moves to the next vial) when FractionCollectionMode is Time. For example, if MaxCollectionPeriod is 120 Second, the fraction collector continues to collect fractions in the next container in line after 120 Second.
Default Calculation: If FractionCollection is True, automatically set according to the MaxCollectionPeriod in the method specified by FractionCollectionMethod option, if available. Otherwise automatically set to the time it takes to fill to the MaxFractionVolume based on the flow rates.
AbsoluteThreshold
The signal value from FractionCollectionDetector above which fractions will always be collected, when FractionCollectionMode is Threshold.
Default Calculation: Inherited from a method specified by FractionCollectionMethod option or set based on FractionCollectionDetector if FractionCollectionMode is Threshold. If the FractionCollectionDetector is UVVis, automatically set to 500 Milli AbsorbanceUnit. If the FractionCollectionDetector is Fluorescence, automatically set to 100 Milli RFU.
Programmatic Pattern: ((RangeP[0, 14] | (GreaterEqualP[0*AbsorbanceUnit] | GreaterEqualP[0*RFU] | GreaterEqualP[10*Micro*(Siemens/Centimeter)])) | Automatic) | Null
PeakSlope
The minimum slope (signal change per second) required for PeakSlopeDuration to trigger peak detection and start fraction collection. Fraction collection end slope is defined as the opposite of PeakSlope and fraction collection will continue until the slope exceeds the negative of the PeakSlope. For instance, if PeakSlope is set to 1 Milli Absorbance Unit/Second, fraction collection begins when the slope surpasses this value and ends when the slope falls below -1 Milli Absorbance Unit/Second. If a PeakEndThreshold is specified, both the PeakEndThreshold and PeakSlope conditions must be satisfied to stop fraction collection. A new peak and corresponding fraction can be registered when the slope exceeds the PeakSlope again.
Default Calculation: If FractionCollection is True, automatically set according to the PeakSlope in the method specified by FractionCollectionMethod option, if available. If the FractionCollectionDetector is UVVis and FractionCollectionMode is Peak, automatically set to 1 Milli AbsorbanceUnit/Second. If the FractionCollectionDetector is Fluorescence and FractionCollectionMode is Peak, automatically set to 0.2 Milli RFU/Second.
Programmatic Pattern: ((GreaterEqualP[0*(1/Second)] | (GreaterEqualP[0*(AbsorbanceUnit/Second)] | GreaterEqualP[0*(RFU/Second)] | GreaterEqualP[10*Micro*(Siemens/Centimeter/Second)])) | Automatic) | Null
PeakSlopeDuration
The minimum duration that changes in slopes must be maintained before fraction collection is registered or ended. This option is only applicable for UltiMate 3000 HPLC instruments.
Default Calculation: If FractionCollection is True, automatically set according to the PeakSlopeDuration in the method specified by FractionCollectionMethod option, if available. Otherwise automatically set to 0.5 Second if FractionCollectionMode is Peak.
Pattern Description: Greater than or equal to 0 seconds and less than or equal to 4 seconds or Null.
PeakEndThreshold
The signal value below which the end of a peak is marked and fraction collection stops when FractionCollectionMode is Peak. Both the PeakEndThreshold and PeakSlope conditions must be satisfied to stop fraction collection. This option is only applicable for UltiMate 3000 HPLC instruments.
Default Calculation: If FractionCollection is True, automatically set according to the PeakEndThreshold in the method specified by FractionCollectionMethod option, if available. If FractionCollectionMode is Peak, automatically set to 1 Milli AbsorbanceUnit for UVVis detector, 0.2 Milli * RFU for Fluorescence detector, 10 for pH detector or 10.0 Milli * Siemens / Centimeter for Conductivity detector.
Programmatic Pattern: ((RangeP[0, 14] | (GreaterEqualP[0*AbsorbanceUnit] | GreaterEqualP[0*RFU] | GreaterEqualP[10*Micro*(Siemens/Centimeter)])) | Automatic) | Null
Standards
Standard
The reference compound(s) to inject to the instrument, often used for quantification or to check internal measurement consistency.
Default Calculation: If any other Standard option is specified, automatically set based on the SeparationMode option. If InjectionTable is specified, set from the specified Standard entries in the InjectionTable.
Pattern Description: An object of type or subtype Model[Sample] or Object[Sample] or a prepared sample or Null.
StandardInjectionVolume
The physical quantity of Standard sample loaded into the flow path on the selected instrument along with the mobile phase onto the stationary phase.
Pattern Description: Greater than or equal to 0 microliters and less than or equal to 16 milliliters or Null.
StandardFrequency
Default Calculation: If injectionTable is given, automatically set to Null and the sequence of Standards specified in the InjectionTable will be used in the experiment. If any other Standard option is specified, automatically set to FirstAndLast.
Pattern Description: Greater than 0 in increments of 1 or None, First, Last, FirstAndLast, or GradientChange or Null.
Programmatic Pattern: (((None | First | Last | FirstAndLast | GradientChange) | GreaterP[0, 1]) | Automatic) | Null
StandardColumnPosition
The position of the column selector valve and the desired column configuration that will be used for each standard sample as it is injected.
Default Calculation: If InjectionTable is specified, automatically set from the Column Position entry for the standard sample. Otherwise set to PositionA.
Pattern Description: PositionA, PositionB, PositionC, PositionD, PositionE, PositionF, PositionG, or PositionH or Null.
StandardColumnTemperature
Default Calculation: Automatically set to the corresponding gradient temperature specified in the StandardGradient option or the column temperature for the sample in the InjectionTable option; otherwise, set as the first value of the ColumnTemperature option.
Pattern Description: Ambient or greater than or equal to 5 degrees Celsius and less than or equal to 90 degrees Celsius or Null.
StandardGradientA
The composition of BufferA within the flow, defined for specific time points for Standard measurement. The composition is linearly interpolated for the intervening periods between the defined time points. For example for StandardGradientA->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferA in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If StandardGradient option is specified, set from it or implicitly determined from the StandardGradientB, StandardGradientC, and StandardGradientD options such that the total amounts to 100%.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or list of one or more {Time, Buffer A Composition} entries or Null.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic) | Null
StandardGradientB
The composition of BufferB within the flow, defined for specific time points for Standard measurement. The composition is linearly interpolated for the intervening periods between the defined time points. For example for StandardGradientB->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferB in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If StandardGradient option is specified, set from it or implicitly determined from the StandardGradientA, StandardGradientC, and StandardGradientD options such that the total amounts to 100%.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or list of one or more {Time, Buffer B Composition} entries or Null.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic) | Null
StandardGradientC
The composition of BufferC within the flow, defined for specific time points for Standard measurement. The composition is linearly interpolated for the intervening periods between the defined time points. For example for StandardGradientC->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferC in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If StandardGradient option is specified, set from it or implicitly determined from the StandardGradientA, StandardGradientB, and StandardGradientD options such that the total amounts to 100%.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or list of one or more {Time, Buffer C Composition} entries or Null.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic) | Null
StandardGradientD
The composition of BufferD within the flow, defined for specific time points for Standard measurement. The composition is linearly interpolated for the intervening periods between the defined time points. For example for StandardGradientD->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferD in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If StandardGradient option is specified, set from it or implicitly determined from the StandardGradientA, StandardGradientB, and StandardGradientC options such that the total amounts to 100%.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or list of one or more {Time, Buffer D Composition} entries or Null.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic) | Null
StandardFlowRate
The net speed of the fluid flowing through the pump inclusive of the composition of BufferA, BufferB, BufferC, and BufferD specified in the StandardGradient options during Standard measurement. This speed is linearly interpolated such that consecutive entries of {Time, Flow Rate} will define the intervening fluid speed. For example, {{0 Minute, 0.3 Milliliter/Minute},{30 Minute, 0.5 Milliliter/Minute}} means flow rate of 0.4 Milliliter/Minute at 15 minutes into the run.
Default Calculation: If StandardGradient option is specified, automatically set from the method given in the StandardGradient option. If NominalFlowRate of the column model is specified, set to lesser of the NominalFlowRate for each of the columns, guard columns or the instrument's MaxFlowRate. Otherwise set to 1 Milliliter / Minute.
Pattern Description: Greater than or equal to 0 milliliters per minute and less than or equal to 200 milliliters per minute or list of one or more {Time, Flow Rate} entries or Null.
Programmatic Pattern: ((RangeP[0*(Milliliter/Minute), 200*(Milliliter/Minute)] | {{GreaterEqualP[0*Minute], RangeP[0*(Milliliter/Minute), 200*(Milliliter/Minute)]}..}) | Automatic) | Null
StandardGradient
The composition of different specified buffers in BufferA, BufferB, BufferC and BufferD over time in the fluid flow for Standard measurement. Specific parameters of a gradient object can be overridden by specific options. Differential Refractive Index Reference Loading refers to the HPLC refractive index loading reference values as shown in the Figure 2.7.4. When open, the flow downstream of the column is loaded into the two flow cells simultaneously.
Default Calculation: Automatically set to best meet all the StandardGradient options (e.g. StandardGradientA, StandardGradientB, StandardGradientC, StandardGradientD, StandardFlowRate).
Pattern Description: An object of type or subtype Object[Method, Gradient] or list of one or more {Time, Buffer A Composition, Buffer B Composition, Buffer C Composition, Buffer D Composition, Flow Rate, Differential Refractive Index Reference Loading} entries or Null.
Programmatic Pattern: ((ObjectP[Object[Method, Gradient]] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent], RangeP[0*Percent, 100*Percent], RangeP[0*Percent, 100*Percent], RangeP[0*Percent, 100*Percent], RangeP[0*(Milliliter/Minute), 200*(Milliliter/Minute)], Open | Closed | None | Automatic}..}) | Automatic) | Null
StandardAbsorbanceWavelength
For Standard measurement, the wavelength of light passed through the flow cell for the UVVis Detector. For PhotoDiodeArray Detector, a 3D data is generated from a spectrum of light passing through the flow cell. Absorbance wavelength in that case represents the wavelength at which a 2D data slice is generated from the 3D data.
Default Calculation: If a UVVis Detector or PhotoDiodeArray Detector is selected or available on the selected instrument, automatically set equal to the same value as the first entry in AbsorbanceWavelength.
Programmatic Pattern: ((RangeP[190*Nanometer, 950*Nanometer] | All | RangeP[190*Nanometer, 950*Nanometer] ;; RangeP[200*Nanometer, 950*Nanometer]) | Automatic) | Null
StandardWavelengthResolution
The increment in wavelength for the range of wavelength of light passed through the flow for absorbance measurement for the instruments with PhotoDiodeArray Detector for Standard measurement.
Default Calculation: If a PhotoDiodeArray Detector is selected or available on the selected Instrument, automatically set equal to the same value as the first entry in WavelengthResolution.
Pattern Description: Greater than or equal to 0.1 nanometers and less than or equal to 12. nanometers or Null.
StandardUVFilter
Indicates if UV wavelengths (less than 210 nm) should be blocked from being transmitted through the sample for the PhotoDiodeArray Detector for Standard measurement.
Default Calculation: If a PhotoDiodeArray Detector is selected or available on the selected Instrument, automatically set to the first entry in UVFilter.
StandardAbsorbanceSamplingRate
The number of times an absorbance measurement is made per second for Standard sample. Lower values will be less susceptible to noise but will record less frequently across time.
Default Calculation: If a UVVis Detector or PhotoDiodeArray Detector is selected or available on the selected instrument, automatically set equal to the first entry in AbsorbanceSamplingRate.
Pattern Description: Greater than or equal to 1 reciprocal second and less than or equal to 120 reciprocal seconds or Null.
StandardExcitationWavelength
The wavelength(s) that is used to excite fluorescence in the Standard sample in the Fluorescence Detector.
Default Calculation: If Fluorescence Detector is selected, automatically set to the first entry in ExcitationWavelength.
Programmatic Pattern: ((RangeP[200*Nanometer, 1200*Nanometer] | {RangeP[200*Nanometer, 890*Nanometer]..}) | Automatic) | Null
StandardEmissionWavelength
The wavelength(s) of light at which fluorescence emitted from the Standard sample is measured in the Fluorescence Detector.
Default Calculation: If Fluorescence Detector is selected, automatically set to the first entry in EmissionWavelength.
Programmatic Pattern: ((RangeP[200*Nanometer, 1200*Nanometer] | {RangeP[200*Nanometer, 890*Nanometer]..}) | Automatic) | Null
StandardEmissionCutOffFilter
The cut-off wavelength to pre-select the emitted light from the Standard sample and allow only the light with wavelength above the desired value to pass, before the light enters emission monochromator for final wavelength selection for Ultimate 3000 with FLR Detector. If set to None, no cut-off filter is used.
Default Calculation: If a Fluorescence Detector with a cut-off filter wheel is selected, automatically set to the first entry in EmissionCutOffFilter.
Pattern Description: 280 nanometers, 370 nanometers, 435 nanometers, 530 nanometers, or None or Null.
StandardFluorescenceGain
For each StandardExcitationWavelength/StandardEmissionWavelength pair, the signal amplification factor which modulates the percentage of maximum voltage that can be applied to the Photomultiplier Tube of the Fluorescence Detector during Standard measurement. Linear increase in voltage applied to the Photomultiplier tube leads to an exponential change in RFU signal. Variable Fluorescence Sensitivity implies a different fluorescence sensitivity for each Excitation/Emission Wavelength pair.
Default Calculation: If Fluorescence Detector is selected, automatically set to the first entry in FluorescenceGain.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {RangeP[0*Percent, 100*Percent]..}) | Automatic) | Null
StandardFluorescenceFlowCellTemperature
The temperature that the thermostat inside the fluorescence flow cell of the Fluorescence Detector is set to during the fluorescence measurement of the Standard sample.
Default Calculation: If Fluorescence Detector is selected and temperature control is available on that unit, automatically set to the first entry in FluorescenceFlowCellTemperature.
Pattern Description: Ambient or greater than or equal to 25 degrees Celsius and less than or equal to 50 degrees Celsius or Null.
StandardLightScatteringLaserPower
The laser power filter used in the Multi-Angle static Light Scattering (MALS) and Dynamic Light Scattering (DLS) Detector for the measurement of the Standard sample. 100% means that no filter is being used to restrict the laser power.
Default Calculation: If MultiAngleLightScattering Detector and/or DynamicLightScattering Detector are selected, automatically set to the first entry in LightScatteringLaserPower.
Pattern Description: Greater than or equal to 10 percent and less than or equal to 100 percent or Null.
StandardLightScatteringFlowCellTemperature
The temperature that the thermostat inside the flow cell of the Detector is set to during the Multi-Angle static Light Scattering (MALS) and/or Dynamic Light Scattering (DLS) measurement of the Standard sample.
Default Calculation: If MultiAngleLightScattering Detector and/or DynamicLightScattering Detector are selected, automatically set to the first entry in LightScatteringFlowCellTemperature.
Pattern Description: Ambient or greater than or equal to -15 degrees Celsius and less than or equal to 210 degrees Celsius or Null.
StandardRefractiveIndexMethod
The type of refractive index measurement of the Refractive Index (RI) Detector for the measurement of the Standard. When DifferentialRefractiveIndex is selected, the refractive index difference between the flow downstream sample and the reference solvent is measured. See Figure 2.7.4 for more information.
Default Calculation: If RefractiveIndex Detector is selected and Differential Refractive Index Reference Loading is set to Closed in StandardGradient, automatically set to DifferentialRefractiveIndex. Otherwise automatically set to RefractiveIndex.
StandardRefractiveIndexFlowCellTemperature
The temperature that the thermostat inside the refractive index flow cell of the Refractive Index (RI) Detector is set to during the refractive index measurement of the Standard sample.
Default Calculation: If RefractiveIndex Detector is selected, automatically set to the first entry in RefractiveIndexFlowCellTemperature.
Pattern Description: Ambient or greater than or equal to 4 degrees Celsius and less than or equal to 65 degrees Celsius or Null.
StandardNebulizerGas
Indicates if Nitrogen sheath gas is flowed along with the Standard sample within the EvaporativeLightScattering Detector.
Default Calculation: If EvaporativeLightScattering is selected, automatically set to the first entry in NebulizerGas.
StandardNebulizerGasHeating
Indicates if the sheath gas that carries samples in the EvaporativeLightScattering Detector is heated for Standard measurement.
Default Calculation: If EvaporativeLightScattering Detector is selected and StandardNebulizerGas is True, automatically set to the first entry in NebulizerGasHeating.
StandardNebulizerHeatingPower
The relative magnitude of the heating element used to heat the sheath gas for the EvaporativeLightScattering Detector for Standard measurement (Corresponding temperature not measured by the manufacturer). Higher percent values correspond to percent of power going to heating coil.
Default Calculation: If EvaporativeLightScattering Detector is selected and StandardNebulizerGas is True, automatically set to the first entry in NebulizerHeatingPower.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or Null.
StandardNebulizerGasPressure
The applied pressure of sheath gas for the EvaporativeLightScattering Detector for Standard measurement (Flow rate unmeasured by the manufacturer). Higher pressure corresponds to faster sheath gas flow.
Default Calculation: If EvaporativeLightScattering Detector is selected and StandardNebulizerGas is True, automatically set to the first entry in NebulizerGasPressure.
Pattern Description: Greater than or equal to 20 pounds‐force per inch squared and less than or equal to 60 pounds‐force per inch squared or Null.
StandardDriftTubeTemperature
The set temperature of the chamber thermostat through which the nebulized analytes flow within the EvaporativeLightScattering Detector for Standard samples. The purpose to heat the drift tube is to evaporate any unevaporated solvent remaining in the flow from the nebulizer.
Default Calculation: If EvaporativeLightScattering Detector is selected and StandardNebulizerGas is True, automatically set to the first entry in DriftTubeTemperature.
Pattern Description: Greater than or equal to 20 degrees Celsius and less than or equal to 100 degrees Celsius or Null.
StandardELSDGain
The percent of maximum voltage sent to the Photo Multiplier Tube (PMT) for signal amplification for the EvaporativeLightScattering measurement. The percentage value specified here is converted into a unitless factor from 0 to 1000 which the software accepts to modulate the voltage for the PMT.
Default Calculation: If EvaporativeLightScattering Detector is selected and StandardNebulizerGas is True, automatically set to the first entry in ELSDGain.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or Null.
StandardELSDSamplingRate
The frequency of evaporative light scattering measurement for Standard samples. Lower values will be less susceptible to noise but will record less frequently across time. Lower or higher values do not affect the y axis of the measurement.
Default Calculation: If EvaporativeLightScattering Detector is selected and NebulizerGas is True, automatically set to the first entry in ELSDSamplingRate.
Pattern Description: Greater than or equal to 1 reciprocal second and less than or equal to 80 reciprocal seconds or Null.
StandardStorageCondition
The non-default conditions under which any standards used by this experiment should be stored after the protocol is completed. If left unset, Standard samples will be stored according to their Models' DefaultStorageCondition.
Pattern Description: {AmbientStorage, Refrigerator, Freezer, DeepFreezer, CryogenicStorage, YeastIncubation, YeastShakingIncubation, BacterialIncubation, BacterialShakingIncubation, MammalianIncubation, ViralIncubation, CrystalIncubation, AcceleratedTesting, IntermediateTesting, LongTermTesting, UVVisLightTesting} or Disposal or Null.
Blanks
Blank
The object(s) (samples) to inject typically as negative controls (e.g. to test effects stemming from injection, sample solvent, impurities on the column or buffer).
Default Calculation: If any other Blank option is specified or RefractiveIndex Detector is selected, automatically set to the specified BufferA or Model[Sample, "Milli-Q water"].
Pattern Description: An object of type or subtype Model[Sample] or Object[Sample] or a prepared sample or Null.
BlankInjectionVolume
The physical quantity of Blank sample that is loaded into the flow path on the selected instrument along with the mobile phase onto the stationary phase.
Pattern Description: Greater than or equal to 0 microliters and less than or equal to 16 milliliters or Null.
BlankFrequency
Default Calculation: If injectionTable is given, automatically set to Null and the sequence of Blanks specified in the InjectionTable will be used in the experiment. If any other Blank option is specified, automatically set to FirstAndLast.
Pattern Description: Greater than 0 in increments of 1 or None, First, Last, FirstAndLast, or GradientChange or Null.
Programmatic Pattern: (((None | First | Last | FirstAndLast | GradientChange) | GreaterP[0, 1]) | Automatic) | Null
BlankColumnPosition
The position of the column selector valve and the desired column configuration that will be used for each blank sample as it is injected.
Default Calculation: For a batch of samples automatically set from the specified Column option. If InjectionTable is specified, set from the Column Position for blank Type injections.
Pattern Description: PositionA, PositionB, PositionC, PositionD, PositionE, PositionF, PositionG, or PositionH or Null.
BlankColumnTemperature
Default Calculation: Automatically set to the corresponding gradient temperature specified in the BlankGradient option or the column temperature for the sample in the InjectionTable option; otherwise, set as the first value of the ColumnTemperature option.
Pattern Description: Ambient or greater than or equal to 5 degrees Celsius and less than or equal to 90 degrees Celsius or Null.
BlankGradientA
The composition of BufferA within the flow, defined for specific time points for Blank measurement. The composition is linearly interpolated for the intervening periods between the defined time points. For example for BlankGradientA->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferA in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If BlankGradient option is specified, set from it or implicitly determined from the BlankGradientB, BlankGradientC, and BlankGradientD options such that the total amounts to 100%.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or list of one or more {Time, Buffer A Composition} entries or Null.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic) | Null
BlankGradientB
The composition of BufferB within the flow, defined for specific time points for Blank measurement. The composition is linearly interpolated for the intervening periods between the defined time points. For example for BlankGradientB->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferB in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If BlankGradient option is specified, set from it or implicitly determined from the BlankGradientA, BlankGradientC, and BlankGradientD options such that the total amounts to 100%.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or list of one or more {Time, Buffer B Composition} entries or Null.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic) | Null
BlankGradientC
The composition of BufferC within the flow, defined for specific time points for Blank measurement. The composition is linearly interpolated for the intervening periods between the defined time points. For example for BlankGradientC->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferC in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If BlankGradient option is specified, set from it or implicitly determined from the BlankGradientA, BlankGradientB, and BlankGradientD options such that the total amounts to 100%.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or list of one or more {Time, Buffer C Composition} entries or Null.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic) | Null
BlankGradientD
The composition of BufferD within the flow, defined for specific time points for Blank measurement. The composition is linearly interpolated for the intervening periods between the defined time points. For example for BlankGradientD->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferD in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If BlankGradient option is specified, set from it or implicitly determined from the BlankGradientA, BlankGradientB, and BlankGradientC options such that the total amounts to 100%.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or list of one or more {Time, Buffer D Composition} entries or Null.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic) | Null
BlankFlowRate
The net speed of the fluid flowing through the pump inclusive of the composition of BufferA, BufferB, BufferC, and BufferD specified in the BlankGradient options during Blank measurement. This speed is linearly interpolated such that consecutive entries of {Time, Flow Rate} will define the intervening fluid speed. For example, {{0 Minute, 0.3 Milliliter/Minute},{30 Minute, 0.5 Milliliter/Minute}} means flow rate of 0.4 Milliliter/Minute at 15 minutes into the run.
Default Calculation: If BlankGradient option is specified, automatically set from the method given in the BlankGradient option. If NominalFlowRate of the column model is specified, set to lesser of the NominalFlowRate for each of the columns, guard columns or the instrument's MaxFlowRate. Otherwise set to 1 Milliliter / Minute.
Pattern Description: Greater than or equal to 0 milliliters per minute and less than or equal to 200 milliliters per minute or list of one or more {Time, Flow Rate} entries or Null.
Programmatic Pattern: ((RangeP[0*(Milliliter/Minute), 200*(Milliliter/Minute)] | {{GreaterEqualP[0*Minute], RangeP[0*(Milliliter/Minute), 200*(Milliliter/Minute)]}..}) | Automatic) | Null
BlankGradient
The composition of different specified buffers in BufferA, BufferB, BufferC and BufferD over time in the fluid flow during Blank measurement. Specific parameters of a gradient object can be overridden by specific options. Differential Refractive Index Reference Loading refers to the HPLC refractive index loading reference values as shown in the Figure 2.7.4. When open, the flow downstream of the column is loaded into the two flow cells simultaneously.
Default Calculation: Automatically set to best meet all the BlankGradient options (e.g. BlankGradientA, BlankGradientB, BlankGradientC, BlankGradientD, BlankFlowRate).
Pattern Description: An object of type or subtype Object[Method, Gradient] or list of one or more {Time, Buffer A Composition, Buffer B Composition, Buffer C Composition, Buffer D Composition, Flow Rate, Differential Refractive Index Reference Loading} entries or Null.
Programmatic Pattern: ((ObjectP[Object[Method, Gradient]] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent], RangeP[0*Percent, 100*Percent], RangeP[0*Percent, 100*Percent], RangeP[0*Percent, 100*Percent], RangeP[0*(Milliliter/Minute), 200*(Milliliter/Minute)], Open | Closed | None | Automatic}..}) | Automatic) | Null
BlankAbsorbanceWavelength
For Blank measurement, the wavelength of light passed through the flow cell for the UVVis Detector. For PhotoDiodeArray Detector, a 3D data is generated from a spectrum of light passing through the flow cell. Absorbance wavelength in that case represents the wavelength at which a 2D data slice is generated from the 3D data.
Default Calculation: If a UVVis Detector or PhotoDiodeArray Detector is selected or available on the selected instrument, automatically set as the first entry in AbsorbanceWavelength.
Programmatic Pattern: ((RangeP[190*Nanometer, 950*Nanometer] | All | RangeP[190*Nanometer, 950*Nanometer] ;; RangeP[200*Nanometer, 950*Nanometer]) | Automatic) | Null
BlankWavelengthResolution
The increment in wavelength for the range of wavelength of light passed through the flow for absorbance measurement for the instruments with PhotoDiodeArray Detector for Blank measurement.
Default Calculation: If a PhotoDiodeArray Detector is selected or available on the selected instrument, automatically set as the first entry in WavelengthResolution.
Pattern Description: Greater than or equal to 0.1 nanometers and less than or equal to 12. nanometers or Null.
BlankUVFilter
Indicates if UV wavelengths (less than 210 nm) should be blocked from being transmitted through the sample for the PhotoDiodeArray Detector for Blank measurement.
Default Calculation: If a PhotoDiodeArray Detector is selected or available on the selected instrument, automatically set as the first entry in UVFilter.
BlankAbsorbanceSamplingRate
The number of times the absorbance measurement is made per second during Blank measurement. Lower values will be less susceptible to noise but will record less frequently across time.
Default Calculation: If a UVVis Detector or PhotoDiodeArray Detector is selected or available on the selected instrument, automatically set equal to the first entry in AbsorbanceSamplingRate.
Pattern Description: Greater than or equal to 1 reciprocal second and less than or equal to 120 reciprocal seconds or Null.
BlankExcitationWavelength
The wavelength(s) that is used to excite fluorescence in the Blank sample in the Fluorescence Detector.
Default Calculation: If Fluorescence Detector is selected, automatically set to the first entry in ExcitationWavelength.
Programmatic Pattern: ((RangeP[200*Nanometer, 1200*Nanometer] | {RangeP[200*Nanometer, 890*Nanometer]..}) | Automatic) | Null
BlankEmissionWavelength
The wavelength(s) of light at which fluorescence emitted from the Blank sample is measured in the Fluorescence Detector.
Default Calculation: If Fluorescence Detector is selected, automatically set to the first entry in EmissionWavelength.
Programmatic Pattern: ((RangeP[200*Nanometer, 1200*Nanometer] | {RangeP[200*Nanometer, 890*Nanometer]..}) | Automatic) | Null
BlankEmissionCutOffFilter
The cut-off wavelength to pre-select the emitted light from the Blank sample and allow only the light with wavelength above the desired value to pass, before the light enters emission monochromator for final wavelength selection for Ultimate 3000 with FLR Detector. If set to None, no cut-off filter is used.
Default Calculation: If a Fluorescence Detector with a cut-off filter wheel is selected, automatically set to the first entry in EmissionCutOffFilter.
Pattern Description: 280 nanometers, 370 nanometers, 435 nanometers, 530 nanometers, or None or Null.
BlankFluorescenceGain
For each BlankExcitationWavelength/BlankEmissionWavelength pair, the signal amplification factor which modulates the percentage of maximum voltage that can be applied to the Photomultiplier Tube of the Fluorescence Detector during Standard measurement. Linear increase in voltage applied to the Photomultiplier tube leads to an exponential change in RFU signal. Variable Fluorescence Sensitivity implies a different fluorescence sensitivity for each Excitation/Emission Wavelength pair.
Default Calculation: If Fluorescence Detector is selected, automatically set to the first entry in FluorescenceGain.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {RangeP[0*Percent, 100*Percent]..}) | Automatic) | Null
BlankFluorescenceFlowCellTemperature
The temperature that the thermostat inside the fluorescence flow cell of the Fluorescence Detector is set to during the fluorescence measurement of the Blank sample.
Default Calculation: If Fluorescence Detector is selected and temperature control is available on that unit, automatically set to the first entry in FluorescenceFlowCellTemperature.
Pattern Description: Ambient or greater than or equal to 25 degrees Celsius and less than or equal to 50 degrees Celsius or Null.
BlankLightScatteringLaserPower
The laser power filter used in the Multi-Angle static Light Scattering (MALS) and Dynamic Light Scattering (DLS) Detector for the measurement of the Blank sample. 100% means that no filter is being used to restrict the laser power.
Default Calculation: If MultiAngleLightScattering Detector and/or DynamicLightScattering Detector are selected, automatically set to the first entry in LightScatteringLaserPower.
Pattern Description: Greater than or equal to 10 percent and less than or equal to 100 percent or Null.
BlankLightScatteringFlowCellTemperature
The temperature that the thermostat inside the flow cell of the Detector is set to during the Multi-Angle static Light Scattering (MALS) and/or Dynamic Light Scattering (DLS) measurement of the Blank sample.
Default Calculation: If MultiAngleLightScattering Detector and/or DynamicLightScattering Detector are selected, automatically set to the first entry in LightScatteringFlowCellTemperature.
Pattern Description: Ambient or greater than or equal to -15 degrees Celsius and less than or equal to 210 degrees Celsius or Null.
BlankRefractiveIndexMethod
The type of refractive index measurement of the Refractive Index (RI) Detector for the measurement of the Blank. When DifferentialRefractiveIndex is selected, the refractive index difference between the flow downstream sample and the reference solvent is measured. See Figure 2.7.4 for more information.
Default Calculation: If RefractiveIndex Detector is selected and Differential Refractive Index Reference Loading is set to Closed in BlankGradient, automatically set to DifferentialRefractiveIndex. Otherwise automatically set to RefractiveIndex.
BlankRefractiveIndexFlowCellTemperature
The temperature that the thermostat inside the refractive index flow cell of the Refractive Index (RI) Detector is set to during the refractive index measurement of the Blank sample.
Default Calculation: If RefractiveIndex Detector is selected, automatically set to the first entry in RefractiveIndexFlowCellTemperature.
Pattern Description: Ambient or greater than or equal to 4 degrees Celsius and less than or equal to 65 degrees Celsius or Null.
BlankNebulizerGas
Indicates if Nitrogen sheath gas is flowed along with the Blank sample within the EvaporativeLightScattering Detector.
Default Calculation: If EvaporativeLightScattering is selected, automatically set to the first entry in NebulizerGas.
BlankNebulizerGasHeating
Indicates if the sheath gas that carries samples in the EvaporativeLightScattering Detector is heated for Blank measurement.
Default Calculation: If EvaporativeLightScattering Detector is selected and BlankNebulizerGas is True, automatically set to the first entry in NebulizerGasHeating.
BlankNebulizerHeatingPower
The relative magnitude of the heating element used to heat the sheath gas for the EvaporativeLightScattering Detector for Blank measurement (Corresponding temperature not measured by the manufacturer). Higher percent values correspond to percent of power going to heating coil.
Default Calculation: If EvaporativeLightScattering Detector is selected and BlankNebulizerGas is True, automatically set to the first entry in NebulizerHeatingPower.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or Null.
BlankNebulizerGasPressure
The applied pressure of sheath gas for the EvaporativeLightScattering Detector for Blank measurement (Flow rate unmeasured by the manufacturer). Higher pressure corresponds to faster sheath gas flow.
Default Calculation: If EvaporativeLightScattering Detector is selected and BlankNebulizerGas is True, automatically set to the first entry in NebulizerGasPressure.
Pattern Description: Greater than or equal to 20 pounds‐force per inch squared and less than or equal to 60 pounds‐force per inch squared or Null.
BlankDriftTubeTemperature
The set temperature of the chamber thermostat through which the nebulized analytes flow within the EvaporativeLightScattering Detector for Blank samples. The purpose to heat the drift tube is to evaporate any unevaporated solvent remaining in the flow from the nebulizer.
Default Calculation: If EvaporativeLightScattering Detector is selected and BlankNebulizerGas is True, automatically set to the first entry in DriftTubeTemperature.
Pattern Description: Greater than or equal to 20 degrees Celsius and less than or equal to 100 degrees Celsius or Null.
BlankELSDGain
The percent of maximum voltage sent to the Photo Multiplier Tube (PMT) for signal amplification for the EvaporativeLightScattering measurement. The percentage value specified here is converted into a unitless factor from 0 to 1000 which the software accepts to modulate the voltage for the PMT.
Default Calculation: If EvaporativeLightScattering Detector is selected and BlankNebulizerGas is True, automatically set to the first entry in ELSDGain.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or Null.
BlankELSDSamplingRate
The frequency of evaporative light scattering measurement for Blank Samples. Lower values will be less susceptible to noise but will record less frequently across time. Lower or higher values do not affect the y axis of the measurement.
Default Calculation: If EvaporativeLightScattering Detector is selected and BlankNebulizerGas is True, automatically set to the first entry in ELSDSamplingRate.
Pattern Description: Greater than or equal to 1 reciprocal second and less than or equal to 80 reciprocal seconds or Null.
BlankStorageCondition
The non-default conditions under which any blanks used by this experiment should be stored after the protocol is completed. If left unset, Blank samples will be stored according to their Models' DefaultStorageCondition.
Pattern Description: {AmbientStorage, Refrigerator, Freezer, DeepFreezer, CryogenicStorage, YeastIncubation, YeastShakingIncubation, BacterialIncubation, BacterialShakingIncubation, MammalianIncubation, ViralIncubation, CrystalIncubation, AcceleratedTesting, IntermediateTesting, LongTermTesting, UVVisLightTesting} or Disposal or Null.
Column Prime
ColumnRefreshFrequency
The frequency of column prime inserted into the order of analyte injections at which solvent is flowed to equilibrate the column in order to remove contaminants and reset the gradient to match the starting percentage of the subsequent injection. An initial column prime and final column flush will be performed unless Null or None is specified. For First, it is performed at the beginning. For Last, it is performed at the end. For FirstAndLast, it is performed both at the beginning and end. For GradientChange, it is performed every time a change in the gradient is encountered for the injections. A Number indicates the number of injections after which it is performed and also in the beginning (eg: for 2, it is performed at the start and after 2nd, 4th, 6th and so on injections).
Default Calculation: Automatically set to Null when InjectionTable option is specified (meaning that this option is inconsequential) or no column is used in the experiment; otherwise, set to GradientChange.
Pattern Description: Greater than 0 in increments of 1 or None, FirstAndLast, First, Last, or GradientChange or Null.
Programmatic Pattern: (((None | FirstAndLast | First | Last | GradientChange) | GreaterP[0, 1]) | Automatic) | Null
ColumnPrimeTemperature
Default Calculation: Automatically set to the corresponding gradient temperature specified in the ColumnPrimeGradient option or the column temperature for the column prime in the InjectionTable option; otherwise, set as the first value of the ColumnTemperature option.
Pattern Description: Ambient or greater than or equal to 5 degrees Celsius and less than or equal to 90 degrees Celsius or Null.
ColumnPrimeGradientA
The composition of BufferA within the flow, defined for specific time points for column prime. The composition is linearly interpolated for the intervening periods between the defined time points. For example for ColumnPrimeGradientA->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferA in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If ColumnPrimeGradient option is specified, set from it or implicitly determined from the ColumnPrimeGradientB, ColumnPrimeGradientC, and ColumnPrimeGradientD options such that the total amounts to 100%.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or list of one or more {Time, Buffer A Composition} entries or Null.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic) | Null
ColumnPrimeGradientB
The composition of BufferB within the flow, defined for specific time points for column prime. The composition is linearly interpolated for the intervening periods between the defined time points. For example for ColumnPrimeGradientB->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferB in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If ColumnPrimeGradient option is specified, set from it or implicitly determined from the ColumnPrimeGradientA, ColumnPrimeGradientC, and ColumnPrimeGradientD options such that the total amounts to 100%.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or list of one or more {Time, Buffer B Composition} entries or Null.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic) | Null
ColumnPrimeGradientC
The composition of BufferC within the flow, defined for specific time points for column prime. The composition is linearly interpolated for the intervening periods between the defined time points. For example for ColumnPrimeGradientC->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferC in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If ColumnPrimeGradient option is specified, set from it or implicitly determined from the ColumnPrimeGradientA, ColumnPrimeGradientB, and ColumnPrimeGradientD options such that the total amounts to 100%.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or list of one or more {Time, Buffer C Composition} entries or Null.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic) | Null
ColumnPrimeGradientD
The composition of BufferD within the flow, defined for specific time points for column prime. The composition is linearly interpolated for the intervening periods between the defined time points. For example for ColumnPrimeGradientD->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferD in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If ColumnPrimeGradient option is specified, set from it or implicitly determined from the ColumnPrimeGradientA, ColumnPrimeGradientB, and ColumnPrimeGradientC options such that the total amounts to 100%.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or list of one or more {Time, Buffer D Composition} entries or Null.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic) | Null
ColumnPrimeFlowRate
The net speed of the fluid flowing through the pump inclusive of the composition of BufferA, BufferB, BufferC, and BufferD specified in the ColumnPrimeGradient options during column prime. This speed is linearly interpolated such that consecutive entries of {Time, Flow Rate} will define the intervening fluid speed. For example, {{0 Minute, 0.3 Milliliter/Minute},{30 Minute, 0.5 Milliliter/Minute}} means flow rate of 0.4 Milliliter/Minute at 15 minutes into the run.
Default Calculation: If ColumnPrimeGradient option is specified, automatically set from the method given in the ColumnPrimeGradient option. If NominalFlowRate of the column model is specified, set to lesser of the NominalFlowRate for each of the columns, guard columns or the instrument's MaxFlowRate. Otherwise set to 1 Milliliter / Minute.
Pattern Description: Greater than or equal to 0 milliliters per minute and less than or equal to 200 milliliters per minute or list of one or more {Time, Flow Rate} entries or Null.
Programmatic Pattern: ((RangeP[0*(Milliliter/Minute), 200*(Milliliter/Minute)] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*(Milliliter/Minute), 200*(Milliliter/Minute)]}..}) | Automatic) | Null
ColumnPrimeGradient
The composition of different specified buffers in BufferA, BufferB, BufferC and BufferD over time in the fluid flow for column prime. Specific parameters of a gradient object can be overridden by specific options. Differential Refractive Index Reference Loading refers to the HPLC refractive index loading reference values as shown in the Figure 2.7.4. When open, the flow downstream of the column is loaded into the two flow cells simultaneously.
Default Calculation: Automatically set to best meet all the ColumnPrimeGradient options (e.g. ColumnPrimeGradientA, ColumnPrimeGradientB, ColumnPrimeGradientC, ColumnPrimeGradientD, ColumnPrimeFlowRate).
Pattern Description: An object of type or subtype Object[Method, Gradient] or list of one or more {Time, Buffer A Composition, Buffer B Composition, Buffer C Composition, Buffer D Composition, Flow Rate, Differential Refractive Index Reference Loading} entries or Null.
Programmatic Pattern: ((ObjectP[Object[Method, Gradient]] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent], RangeP[0*Percent, 100*Percent], RangeP[0*Percent, 100*Percent], RangeP[0*Percent, 100*Percent], RangeP[0*(Milliliter/Minute), 200*(Milliliter/Minute)], Open | Closed | None | Automatic}..}) | Automatic) | Null
ColumnPrimeAbsorbanceWavelength
The wavelength of light passed through the flow for the UVVis and Detector for detection during column prime. For PhotoDiodeArray Detector, a 3D data is generated from a spectrum of light passing through the flow cell. Absorbance wavelength in that case represents the wavelength at which a 2D data slice is generated from the 3D data.
Default Calculation: If a UVVis Detector or PhotoDiodeArray Detector is selected or available on the selected instrument, automatically set equal to the same value as the first entry in AbsorbanceWavelength.
Programmatic Pattern: ((RangeP[190*Nanometer, 950*Nanometer] | All | RangeP[190*Nanometer, 950*Nanometer] ;; RangeP[200*Nanometer, 950*Nanometer]) | Automatic) | Null
ColumnPrimeWavelengthResolution
The increment in wavelength for the range of wavelength of light passed through the flow for absorbance measurement for the instruments with PhotoDiodeArray Detector during column prime.
Default Calculation: If a PhotoDiodeArray Detector is selected or available on the selected instrument, automatically set equal to the same value as the first entry in WavelengthResolution.
Pattern Description: Greater than or equal to 0.1 nanometers and less than or equal to 12. nanometers or Null.
ColumnPrimeUVFilter
Indicates if UV wavelengths (less than 210 nm) should be blocked from being transmitted through the flow for PhotoDiodeArray Detector during column prime.
Default Calculation: If a PhotoDiodeArray Detector is selected or available on the selected instrument, automatically set to the first entry in UVFilter.
ColumnPrimeAbsorbanceSamplingRate
The number of times an absorbance measurement is made per second during column prime. Lower values will be less susceptible to noise but will record less frequently across time.
Default Calculation: If a UVVis Detector or PhotoDiodeArray Detector is selected or available on the selected instrument, automatically set equal to the first entry in AbsorbanceSamplingRate.
Pattern Description: Greater than or equal to 1 reciprocal second and less than or equal to 120 reciprocal seconds or Null.
ColumnPrimeExcitationWavelength
The wavelength(s) of light that is used to excite fluorescence in the Fluorescence Detector during column prime.
Default Calculation: If Fluorescence Detector is selected, automatically set to the first entry in ExcitationWavelength.
Programmatic Pattern: ((RangeP[200*Nanometer, 1200*Nanometer] | {RangeP[200*Nanometer, 890*Nanometer]..}) | Automatic) | Null
ColumnPrimeEmissionWavelength
The wavelength(s) of light at which fluorescence emitted from the flow downstream of the column is measured in the Fluorescence Detector during column prime.
Default Calculation: If Fluorescence Detector is selected, automatically set to the first entry in EmissionWavelength.
Programmatic Pattern: ((RangeP[200*Nanometer, 1200*Nanometer] | {RangeP[200*Nanometer, 890*Nanometer]..}) | Automatic) | Null
ColumnPrimeEmissionCutOffFilter
The cut-off wavelength to pre-select the emitted light from the flow downstream of the column and allow only the light with wavelength above the desired value to pass, before the light enters emission monochromator for final wavelength selection during column prime for Ultimate 3000 with FLR Detector. If set to None, no cut-off filter is used.
Default Calculation: If a Fluorescence Detector with a cut-off filter wheel is selected, automatically set to the first entry in EmissionCutOffFilter.
Pattern Description: 280 nanometers, 370 nanometers, 435 nanometers, 530 nanometers, or None or Null.
ColumnPrimeFluorescenceGain
For each ColumnPrimeExcitationWavelength/ColumnPrimeEmissionWavelength pair, the signal amplification factor which modulates the percentage of maximum voltage that can be applied to the Photomultiplier Tube of the Fluorescence Detector during column prime. Linear increase in voltage applied to the Photomultiplier tube leads to an exponential change in RFU signal. Variable Fluorescence Sensitivity implies a different fluorescence sensitivity for each Excitation/Emission Wavelength pair.
Default Calculation: If Fluorescence Detector is selected, automatically set to the first entry in FluorescenceGain.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {RangeP[0*Percent, 100*Percent]..}) | Automatic) | Null
ColumnPrimeFluorescenceFlowCellTemperature
The temperature that the thermostat inside the fluorescence flow cell of the Fluorescence Detector is set to during column prime.
Default Calculation: If Fluorescence Detector is selected and temperature control is available on that unit, automatically set to the first entry in FluorescenceFlowCellTemperature.
Pattern Description: Ambient or greater than or equal to 25 degrees Celsius and less than or equal to 50 degrees Celsius or Null.
ColumnPrimeLightScatteringLaserPower
The laser power filter used in the Multi-Angle static Light Scattering (MALS) and Dynamic Light Scattering (DLS) Detector during column prime measurement. 100% means that no filter is being used to restrict the laser power.
Default Calculation: If MultiAngleLightScattering Detector and/or DynamicLightScattering Detector are selected, automatically set to the first entry in LightScatteringLaserPower.
Pattern Description: Greater than or equal to 10 percent and less than or equal to 100 percent or Null.
ColumnPrimeLightScatteringFlowCellTemperature
The temperature that the thermostat inside the flow cell inside the Multi-Angle static Light Scattering (MALS) and Dynamic Light Scattering (DLS) detector is set to during column prime.
Default Calculation: If MultiAngleLightScattering Detector and/or DynamicLightScattering Detector are selected, automatically set to the first entry in LightScatteringFlowCellTemperature.
Pattern Description: Ambient or greater than or equal to -15 degrees Celsius and less than or equal to 210 degrees Celsius or Null.
ColumnPrimeRefractiveIndexMethod
The type of refractive index measurement of the Refractive Index (RI) Detector during column prime. When DifferentialRefractiveIndex is selected, the refractive index difference between the flow downstream sample and the reference solvent is measured. See Figure 2.7.4 for more information.
Default Calculation: If RefractiveIndex Detector is selected and Differential Refractive Index Reference Loading is set to Closed in ColumnPrimeGradient, automatically set to DifferentialRefractiveIndex. Otherwise automatically set to RefractiveIndex.
ColumnPrimeRefractiveIndexFlowCellTemperature
The temperature that the thermostat inside the refractive index flow cell of the Refractive Index (RI) Detector is set to during column prime.
Default Calculation: If RefractiveIndex Detector is selected, automatically set to the first entry in RefractiveIndexFlowCellTemperature.
Pattern Description: Ambient or greater than or equal to 4 degrees Celsius and less than or equal to 65 degrees Celsius or Null.
ColumnPrimeNebulizerGas
Indicates if Nitrogen sheath gas is flowed with the buffer(s) within the EvaporativeLightScattering Detector during column prime.
Default Calculation: If EvaporativeLightScattering is selected, automatically set to the first entry in NebulizerGas.
ColumnPrimeNebulizerGasHeating
Indicates if the sheath gas that carries buffer(s) in the EvaporativeLightScattering Detector is heated during column prime.
Default Calculation: If EvaporativeLightScattering Detector is selected and ColumnPrimeNebulizerGas is True, automatically set to the first entry in NebulizerGasHeating.
ColumnPrimeNebulizerHeatingPower
The relative magnitude of the heating element used to heat the sheath gas for the EvaporativeLightScattering Detector during column prime (Corresponding temperature not measured by the manufacturer). Higher percent values correspond to percent of power going to heating coil.
Default Calculation: If EvaporativeLightScattering Detector is selected and ColumnPrimeNebulizerGas is True, automatically set to the first entry in NebulizerHeatingPower.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or Null.
ColumnPrimeNebulizerGasPressure
The applied pressure of sheath gas for the EvaporativeLightScattering Detector during column prime (Flow rate unmeasured by the manufacturer). Higher pressure corresponds to faster sheath gas flow.
Default Calculation: If EvaporativeLightScattering Detector is selected and ColumnPrimeNebulizerGas is True, automatically set to the first entry in NebulizerGasPressure.
Pattern Description: Greater than or equal to 20 pounds‐force per inch squared and less than or equal to 60 pounds‐force per inch squared or Null.
ColumnPrimeDriftTubeTemperature
The set temperature of the chamber thermostat through which the nebulized analytes flow within the EvaporativeLightScattering Detector during Column Prime. The purpose to heat the drift tube is to evaporate any unevaporated solvent remaining in the flow from the nebulizer.
Default Calculation: If EvaporativeLightScattering Detector is selected and ColumnPrimeNebulizerGas is True, automatically set to the first entry in DriftTubeTemperature.
Pattern Description: Greater than or equal to 20 degrees Celsius and less than or equal to 100 degrees Celsius or Null.
ColumnPrimeELSDGain
The percent of maximum voltage sent to the Photo Multiplier Tube (PMT) for signal amplification for the EvaporativeLightScattering measurement. The percentage value specified here is converted into a unitless factor from 0 to 1000 which the software accepts to modulate the voltage for the PMT.
Default Calculation: If EvaporativeLightScattering Detector is selected and ColumnPrimeNebulizerGas is True, automatically set to the first entry in ELSDGain.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or Null.
ColumnPrimeELSDSamplingRate
The frequency of evaporative light scattering measurement during column prime. Lower values will be less susceptible to noise but will record less frequently across time. Lower or higher values do not affect the y axis of the measurement.
Default Calculation: If EvaporativeLightScattering Detector is selected and ColumnPrimeNebulizerGas is True, automatically set to the first entry in ELSDSamplingRate.
Pattern Description: Greater than or equal to 1 reciprocal second and less than or equal to 80 reciprocal seconds or Null.
Column Flush
ColumnFlushTemperature
Default Calculation: Automatically set to the corresponding gradient temperature specified in the ColumnFlushGradient option or the column temperature for the column flush in the InjectionTable option; otherwise, set as the first value of the ColumnTemperature option.
Pattern Description: Ambient or greater than or equal to 5 degrees Celsius and less than or equal to 90 degrees Celsius or Null.
ColumnFlushGradientA
The composition of BufferA within the flow, defined for specific time points for column flush. The composition is linearly interpolated for the intervening periods between the defined time points. For example for ColumnFlushGradientA->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferA in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If ColumnFlushGradient option is specified, set from it or implicitly determined from the ColumnFlushGradientB, ColumnFlushGradientC, and ColumnFlushGradientD options such that the total amounts to 100%.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or list of one or more {Time, Buffer A Composition} entries or Null.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic) | Null
ColumnFlushGradientB
The composition of BufferB within the flow, defined for specific time points for column flush. The composition is linearly interpolated for the intervening periods between the defined time points. For example for ColumnFlushGradientB->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferB in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If ColumnFlushGradient option is specified, set from it or implicitly determined from the ColumnFlushGradientA, ColumnFlushGradientC, and ColumnFlushGradientD options such that the total amounts to 100%.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or list of one or more {Time, Buffer B Composition} entries or Null.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic) | Null
ColumnFlushGradientC
The composition of BufferC within the flow, defined for specific time points for column flush. The composition is linearly interpolated for the intervening periods between the defined time points. For example for ColumnFlushGradientC->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferC in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If ColumnFlushGradient option is specified, set from it or implicitly determined from the ColumnFlushGradientA, ColumnFlushGradientB, and ColumnFlushGradientD options such that the total amounts to 100%.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or list of one or more {Time, Buffer C Composition} entries or Null.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic) | Null
ColumnFlushGradientD
The composition of BufferD within the flow, defined for specific time points for column flush. The composition is linearly interpolated for the intervening periods between the defined time points. For example for ColumnFlushGradientD->{{0 Minute, 10 Percent},{30 Minute, 90 Percent}}, the percentage of BufferD in the flow will rise such that at 15 minutes, the composition should be 50 Percent.
Default Calculation: If ColumnFlushGradient option is specified, set from it or implicitly determined from the ColumnFlushGradientA, ColumnFlushGradientB, and ColumnFlushGradientC options such that the total amounts to 100%.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or list of one or more {Time, Buffer D Composition} entries or Null.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent]}..}) | Automatic) | Null
ColumnFlushFlowRate
The net speed of the fluid flowing through the pump inclusive of the composition of BufferA, BufferB, BufferC, and BufferD specified in the ColumnFlushGradient options during column flush. This speed is linearly interpolated such that consecutive entries of {Time, Flow Rate} will define the intervening fluid speed. For example, {{0 Minute, 0.3 Milliliter/Minute},{30 Minute, 0.5 Milliliter/Minute}} means flow rate of 0.4 Milliliter/Minute at 15 minutes into the run.
Default Calculation: If ColumnFlushGradient option is specified, automatically set from the method given in the ColumnFlushGradient option. If NominalFlowRate of the column model is specified, set to lesser of the NominalFlowRate for each of the columns, guard columns or the instrument's MaxFlowRate. Otherwise set to 1 Milliliter / Minute.
Pattern Description: Greater than or equal to 0 milliliters per minute and less than or equal to 200 milliliters per minute or list of one or more {Time, Flow Rate} entries or Null.
Programmatic Pattern: ((RangeP[0*(Milliliter/Minute), 200*(Milliliter/Minute)] | {{GreaterEqualP[0*Minute], RangeP[0*(Milliliter/Minute), 200*(Milliliter/Minute)]}..}) | Automatic) | Null
ColumnFlushGradient
The composition of different specified buffers in BufferA, BufferB, BufferC and BufferD over time in the fluid flow for column prime. Specific parameters of a gradient object can be overridden by specific options. Differential Refractive Index Reference Loading refers to the HPLC refractive index loading reference values as shown in the Figure 2.7.4. When open, the flow downstream of the column is loaded into the two flow cells simultaneously.
Default Calculation: Automatically set to best meet the values specified in ColumnFlushGradient options (e.g. ColumnFlushGradientA, ColumnFlushGradientB, ColumnFlushGradientC, ColumnFlushGradientD, ColumnFlushFlowRate).
Pattern Description: An object of type or subtype Object[Method, Gradient] or list of one or more {Time, Buffer A Composition, Buffer B Composition, Buffer C Composition, Buffer D Composition, Flow Rate, Differential Refractive Index Reference Loading} entries or Null.
Programmatic Pattern: ((ObjectP[Object[Method, Gradient]] | {{RangeP[0*Minute, $MaxExperimentTime], RangeP[0*Percent, 100*Percent], RangeP[0*Percent, 100*Percent], RangeP[0*Percent, 100*Percent], RangeP[0*Percent, 100*Percent], RangeP[0*(Milliliter/Minute), 200*(Milliliter/Minute)], Open | Closed | None | Automatic}..}) | Automatic) | Null
ColumnFlushAbsorbanceWavelength
The wavelength of light passed through the flow for the UVVis and Detector for detection during column flush. For PhotoDiodeArray Detector, a 3D data is generated from a spectrum of light passing through the flow cell. Absorbance wavelength in that case represents the wavelength at which a 2D data slice is generated from the 3D data.
Default Calculation: If a UVVis Detector or PhotoDiodeArray Detector is selected or available on the selected instrument, automatically set equal to the same value as the first entry in AbsorbanceWavelength.
Programmatic Pattern: ((RangeP[190*Nanometer, 950*Nanometer] | All | RangeP[190*Nanometer, 950*Nanometer] ;; RangeP[200*Nanometer, 950*Nanometer]) | Automatic) | Null
ColumnFlushWavelengthResolution
The increment of wavelength for the range of wavelength of light passed through the flow for absorbance measurement for the instruments with PhotoDiodeArray Detector during column flush.
Default Calculation: If a PhotoDiodeArray Detector is selected or available on the selected instrument, automatically set equal to the same value as the first entry in WavelengthResolution.
Pattern Description: Greater than or equal to 0.1 nanometers and less than or equal to 12. nanometers or Null.
ColumnFlushUVFilter
Indicates if UV wavelengths (less than 210 nm) should be blocked from being transmitted through the flow for PhotoDiodeArray Detector during column flush.
Default Calculation: If a PhotoDiodeArray Detector is selected or available on the selected instrument, automatically set to the first entry in UVFilter.
ColumnFlushAbsorbanceSamplingRate
The number of times an absorbance measurement is made per second during column flush. Lower values will be less susceptible to noise but will record less frequently across time.
Default Calculation: If a UVVis Detector or PhotoDiodeArray Detector is selected or available on the selected instrument, automatically set equal to the first entry in AbsorbanceSamplingRate.
Pattern Description: Greater than or equal to 1 reciprocal second and less than or equal to 120 reciprocal seconds or Null.
ColumnFlushExcitationWavelength
The wavelength(s) of light that is used to excite fluorescence in the Fluorescence Detector during column flush.
Default Calculation: If Fluorescence Detector is selected, automatically set to the first entry in ExcitationWavelength.
Programmatic Pattern: ((RangeP[200*Nanometer, 1200*Nanometer] | {RangeP[200*Nanometer, 890*Nanometer]..}) | Automatic) | Null
ColumnFlushEmissionWavelength
The wavelength(s) of light at which fluorescence emitted from the flow downstream of the column is measured in the Fluorescence Detector during column flush.
Default Calculation: If Fluorescence Detector is selected, automatically set to the first entry in EmissionWavelength.
Programmatic Pattern: ((RangeP[200*Nanometer, 1200*Nanometer] | {RangeP[200*Nanometer, 890*Nanometer]..}) | Automatic) | Null
ColumnFlushEmissionCutOffFilter
The cut-off wavelength to pre-select the emitted light from the flow downstream of the column and allow only the light with wavelength above the desired value to pass, before the light enters emission monochromator for final wavelength selection during column flush for Ultimate 3000 with FLR Detector. If set to None, no cut-off filter is used.
Default Calculation: If a Fluorescence Detector with a cut-off filter wheel is selected, automatically set to the first entry in EmissionCutOffFilter.
Pattern Description: 280 nanometers, 370 nanometers, 435 nanometers, 530 nanometers, or None or Null.
ColumnFlushFluorescenceGain
For each ColumnFlushExcitationWavelength/ColumnFlushEmissionWavelength pair, the signal amplification factor which modulates the percentage of maximum voltage that can be applied to the Photomultiplier Tube of the Fluorescence Detector during column flush. Linear increase in voltage applied to the Photomultiplier tube leads to an exponential change in RFU signal. Variable Fluorescence Sensitivity implies a different fluorescence sensitivity for each Excitation/Emission Wavelength pair.
Default Calculation: If Fluorescence Detector is selected, automatically set to the first entry in FluorescenceGain.
Programmatic Pattern: ((RangeP[0*Percent, 100*Percent] | {RangeP[0*Percent, 100*Percent]..}) | Automatic) | Null
ColumnFlushFluorescenceFlowCellTemperature
The temperature that the thermostat inside the fluorescence flow cell of the Fluorescence Detector is set to during column flush.
Default Calculation: If Fluorescence Detector is selected and temperature control is available on that unit, automatically set to the first entry in FluorescenceFlowCellTemperature.
Pattern Description: Ambient or greater than or equal to 25 degrees Celsius and less than or equal to 50 degrees Celsius or Null.
ColumnFlushLightScatteringLaserPower
The laser power filter used in the Multi-Angle static Light Scattering (MALS) and Dynamic Light Scattering (DLS) Detector during column flush measurement. 100% means that no filter is being used to restrict the laser power.
Default Calculation: If MultiAngleLightScattering Detector and/or DynamicLightScattering Detector are selected, automatically set to the first entry in LightScatteringLaserPower.
Pattern Description: Greater than or equal to 10 percent and less than or equal to 100 percent or Null.
ColumnFlushLightScatteringFlowCellTemperature
The temperature that the thermostat inside the flow cell inside the Multi-Angle static Light Scattering (MALS) and Dynamic Light Scattering (DLS) detector is set to during column flush.
Default Calculation: If MultiAngleLightScattering detector and/or DynamicLightScattering detector are selected, automatically set to the first entry in LightScatteringFlowCellTemperature.
Pattern Description: Ambient or greater than or equal to -15 degrees Celsius and less than or equal to 210 degrees Celsius or Null.
ColumnFlushRefractiveIndexMethod
The type of refractive index measurement of the Refractive Index (RI) Detector during column flush. When DifferentialRefractiveIndex is selected, the refractive index difference between the flow downstream sample and the reference solvent is measured. See Figure 2.7.4 for more information.
Default Calculation: If RefractiveIndex Detector is selected and Differential Refractive Index Reference Loading is set to Closed in ColumnFlushGradient, automatically set to DifferentialRefractiveIndex. Otherwise automatically set to RefractiveIndex.
ColumnFlushRefractiveIndexFlowCellTemperature
The temperature that the thermostat inside the refractive index flow cell of the Refractive Index (RI) Detector is set to during column flush.
Default Calculation: If RefractiveIndex detector is selected, automatically set to the first entry in RefractiveIndexFlowCellTemperature.
Pattern Description: Ambient or greater than or equal to 4 degrees Celsius and less than or equal to 65 degrees Celsius or Null.
ColumnFlushNebulizerGas
Indicates if Nitrogen sheath gas is flowed with the buffer(s) within the EvaporativeLightScattering Detector during column flush.
Default Calculation: If EvaporativeLightScattering is selected, automatically set to the first entry in NebulizerGas.
ColumnFlushNebulizerGasHeating
Indicates if the sheath gas that carries buffer(s) in the EvaporativeLightScattering Detector is heated during column flush.
Default Calculation: If EvaporativeLightScattering Detector is selected and ColumnFlushNebulizerGas is True, automatically set to the first entry in NebulizerGasHeating.
ColumnFlushNebulizerHeatingPower
The relative magnitude of the heating element used to heat the sheath gas for the EvaporativeLightScattering Detector during column flush (Corresponding temperature not measured by the manufacturer). Higher percent values correspond to percent of power going to heating coil.
Default Calculation: If EvaporativeLightScattering Detector is selected and ColumnFlushNebulizerGas is True, automatically set to the first entry in NebulizerHeatingPower.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or Null.
ColumnFlushNebulizerGasPressure
The applied pressure of sheath gas for the EvaporativeLightScattering Detector during column flush (Flow rate unmeasured by the manufacturer). Higher pressure corresponds to faster sheath gas flow.
Default Calculation: If EvaporativeLightScattering Detector is selected and ColumnFlushNebulizerGas is True, automatically set to the first entry in NebulizerGasPressure.
Pattern Description: Greater than or equal to 20 pounds‐force per inch squared and less than or equal to 60 pounds‐force per inch squared or Null.
ColumnFlushDriftTubeTemperature
The set temperature of the chamber thermostat through which the nebulized analytes flow within the EvaporativeLightScattering Detector during Column Flush. The purpose to heat the drift tube is to evaporate any unevaporated solvent remaining in the flow from the nebulizer.
Default Calculation: If EvaporativeLightScattering Detector is selected and ColumnFlushNebulizerGas is True, automatically set to the first entry in DriftTubeTemperature.
Pattern Description: Greater than or equal to 20 degrees Celsius and less than or equal to 100 degrees Celsius or Null.
ColumnFlushELSDGain
The percent of maximum voltage sent to the Photo Multiplier Tube (PMT) for signal amplification for the EvaporativeLightScattering measurement. The percentage value specified here is converted into a unitless factor from 0 to 1000 which the software accepts to modulate the voltage for the PMT.
Default Calculation: If EvaporativeLightScattering Detector is selected and ColumnFlushNebulizerGas is True, automatically set to the first entry in ELSDGain.
Pattern Description: Greater than or equal to 0 percent and less than or equal to 100 percent or Null.
ColumnFlushELSDSamplingRate
The frequency of evaporative light scattering measurement during column flush. Lower values will be less susceptible to noise but will record less frequently across time. Lower or higher values do not affect the y axis of the measurement.
Default Calculation: If EvaporativeLightScattering Detector is selected and ColumnFlushNebulizerGas is True, automatically set to the first entry in ELSDSamplingRate.
Pattern Description: Greater than or equal to 1 reciprocal second and less than or equal to 80 reciprocal seconds or Null.
Sample Preparation
PreparedModelAmount
Indicates the amount of a Model[Sample] specified as input to the experiment function that will be prepared in the PreparedModelContainer.
Programmatic Pattern: RangeP[1*Microliter, 20*Liter] | RangeP[1*Milligram, 20*Kilogram] | GreaterP[0*Unit, 1*Unit] | GreaterP[0., 1.]
PreparedModelContainer
Indicates the container in which a Model[Sample] specified as input to the experiment function will be prepared.
Post Experiment
InjectionSampleVolumeMeasurement
Indicates if any liquid samples prepared in the sub SamplePreparationProtocols of are volume checked prior to injection to provide QC data.
SamplesInStorageCondition
The non-default conditions under which the SamplesIn of this experiment should be stored after the protocol is completed. If left unset, SamplesIn will be stored according to their current StorageCondition.
Pattern Description: {AmbientStorage, Refrigerator, Freezer, DeepFreezer, CryogenicStorage, YeastIncubation, YeastShakingIncubation, BacterialIncubation, BacterialShakingIncubation, MammalianIncubation, ViralIncubation, CrystalIncubation, AcceleratedTesting, IntermediateTesting, LongTermTesting, UVVisLightTesting} or Disposal or Null.
SamplesOutStorageCondition
The non-default conditions under which any new samples generated by this experiment should be stored after the protocol is completed. If left unset, the new samples will be stored according to their Models' DefaultStorageCondition.
Pattern Description: {AmbientStorage, Refrigerator, Freezer, DeepFreezer, CryogenicStorage, YeastIncubation, YeastShakingIncubation, BacterialIncubation, BacterialShakingIncubation, MammalianIncubation, ViralIncubation, CrystalIncubation, AcceleratedTesting, IntermediateTesting, LongTermTesting, UVVisLightTesting} or Disposal or Null.
Sample Prep Options
Sample Preparation
PreparatoryUnitOperations
Specifies a sequence of transferring, aliquoting, consolidating, or mixing of new or existing samples before the main experiment. These prepared samples can be used in the main experiment by referencing their defined name. For more information, please reference the documentation for ExperimentSampleManipulation.
Pattern Description: List of one or more unit Operation ManualSamplePreparation or RoboticSamplePreparation or unit Operation must match SamplePreparationP entries or Null.
Programmatic Pattern: {((ManualSamplePreparationMethodP | RoboticSamplePreparationMethodP) | SamplePreparationP)..} | Null
PreparatoryPrimitives
Specifies a sequence of transferring, aliquoting, consolidating, or mixing of new or existing samples before the main experiment. These prepared samples can be used in the main experiment by referencing their defined name. For more information, please reference the documentation for ExperimentSampleManipulation.
Pattern Description: List of one or more a primitive with head Define, Transfer, Mix, Aliquot, Consolidation, FillToVolume, Incubate, Filter, Wait, Centrifuge, or Resuspend entries or Null.
Preparatory Incubation
Incubate
Indicates if the SamplesIn should be incubated at a fixed temperature prior to starting the experiment or any aliquoting. Sample Preparation occurs in the order of Incubation, Centrifugation, Filtration, and then Aliquoting (if specified).
Default Calculation: Resolves to True if any of the corresponding Incubation options are set. Otherwise, resolves to False.
IncubationTemperature
Temperature at which the SamplesIn should be incubated for the duration of the IncubationTime prior to starting the experiment.
Pattern Description: Ambient or greater than or equal to -20 degrees Celsius and less than or equal to 500 degrees Celsius or Null.
Programmatic Pattern: ((Ambient | RangeP[$MinIncubationTemperature, $MaxIncubationTemperature]) | Automatic) | Null
IncubationTime
Duration for which SamplesIn should be incubated at the IncubationTemperature, prior to starting the experiment.
Mix
Default Calculation: Automatically resolves to True if any Mix related options are set. Otherwise, resolves to False.
MixType
Default Calculation: Automatically resolves based on the container of the sample and the Mix option.
Pattern Description: Roll, Vortex, Sonicate, Pipette, Invert, Stir, Shake, Homogenize, Swirl, Disrupt, or Nutate or Null.
MixUntilDissolved
Indicates if the mix should be continued up to the MaxIncubationTime or MaxNumberOfMixes (chosen according to the mix Type), in an attempt dissolve any solute. Any mixing/incubation will occur prior to starting the experiment.
Default Calculation: Automatically resolves to True if MaxIncubationTime or MaxNumberOfMixes is set.
MaxIncubationTime
Maximum duration of time for which the samples will be mixed while incubated in an attempt to dissolve any solute, if the MixUntilDissolved option is chosen. This occurs prior to starting the experiment.
Default Calculation: Automatically resolves based on MixType, MixUntilDissolved, and the container of the given sample.
IncubationInstrument
Default Calculation: Automatically resolves based on the options Mix, Temperature, MixType and container of the sample.
Pattern Description: An object of type or subtype Model[Instrument, Roller], Model[Instrument, OverheadStirrer], Model[Instrument, Vortex], Model[Instrument, Shaker], Model[Instrument, BottleRoller], Model[Instrument, Roller], Model[Instrument, Sonicator], Model[Instrument, HeatBlock], Model[Instrument, Homogenizer], Model[Instrument, Disruptor], Model[Instrument, Nutator], Model[Instrument, Thermocycler], Model[Instrument, EnvironmentalChamber], Model[Instrument, Pipette], Object[Instrument, Roller], Object[Instrument, OverheadStirrer], Object[Instrument, Vortex], Object[Instrument, Shaker], Object[Instrument, BottleRoller], Object[Instrument, Roller], Object[Instrument, Sonicator], Object[Instrument, HeatBlock], Object[Instrument, Homogenizer], Object[Instrument, Disruptor], Object[Instrument, Nutator], Object[Instrument, Thermocycler], Object[Instrument, EnvironmentalChamber], or Object[Instrument, Pipette] or Null.
AnnealingTime
Minimum duration for which the SamplesIn should remain in the incubator allowing the system to settle to room temperature after the IncubationTime has passed but prior to starting the experiment.
IncubateAliquotContainer
The desired type of container that should be used to prepare and house the incubation samples which should be used in lieu of the SamplesIn for the experiment.
Programmatic Pattern: ((ObjectP[Model[Container]] | {GreaterEqualP[1, 1] | (Automatic | Null), (ObjectP[{Model[Container], Object[Container]}] | _String) | Automatic}) | Automatic) | Null
IncubateAliquotDestinationWell
The desired position in the corresponding IncubateAliquotContainer in which the aliquot samples will be placed.
Default Calculation: Automatically resolves to A1 in containers with only one position. For plates, fills wells in the order provided by the function AllWells.
IncubateAliquot
The amount of each sample that should be transferred from the SamplesIn into the IncubateAliquotContainer when performing an aliquot before incubation.
Default Calculation: Automatically set as the smaller between the current sample volume and the maximum volume of the destination container.
Pattern Description: All or greater than or equal to 1 microliter and less than or equal to 20 liters or Null.
Preparatory Centrifugation
Centrifuge
Indicates if the SamplesIn should be centrifuged prior to starting the experiment or any aliquoting. Sample Preparation occurs in the order of Incubation, Centrifugation, Filtration, and then Aliquoting (if specified).
Default Calculation: Resolves to True if any of the corresponding Centrifuge options are set. Otherwise, resolves to False.
CentrifugeInstrument
Pattern Description: An object of type or subtype Model[Instrument, Centrifuge] or Object[Instrument, Centrifuge] or Null.
Programmatic Pattern: (ObjectP[{Model[Instrument, Centrifuge], Object[Instrument, Centrifuge]}] | Automatic) | Null
CentrifugeIntensity
The rotational speed or the force that will be applied to the samples by centrifugation prior to starting the experiment.
Pattern Description: Greater than 0 revolutions per minute or greater than 0 standard accelerations due to gravity on the surface of the earth or Null.
Programmatic Pattern: ((GreaterP[0*RPM] | GreaterP[0*GravitationalAcceleration]) | Automatic) | Null
CentrifugeTime
CentrifugeTemperature
The temperature at which the centrifuge chamber should be held while the samples are being centrifuged prior to starting the experiment.
Pattern Description: Ambient or greater than or equal to -10 degrees Celsius and less than or equal to 40 degrees Celsius or Null.
CentrifugeAliquotContainer
The desired type of container that should be used to prepare and house the centrifuge samples which should be used in lieu of the SamplesIn for the experiment.
Programmatic Pattern: ((ObjectP[Model[Container]] | {GreaterEqualP[1, 1] | (Automatic | Null), (ObjectP[{Model[Container], Object[Container]}] | _String) | Automatic}) | Automatic) | Null
CentrifugeAliquotDestinationWell
The desired position in the corresponding AliquotContainer in which the aliquot samples will be placed.
Default Calculation: Automatically resolves to A1 in containers with only one position. For plates, fills wells in the order provided by the function AllWells.
CentrifugeAliquot
The amount of each sample that should be transferred from the SamplesIn into the CentrifugeAliquotContainer when performing an aliquot before centrifugation.
Default Calculation: Automatically set as the smaller between the current sample volume and the maximum volume of the destination container.
Pattern Description: All or greater than or equal to 1 microliter and less than or equal to 20 liters or Null.
Preparatory Filtering
Filtration
Indicates if the SamplesIn should be filter prior to starting the experiment or any aliquoting. Sample Preparation occurs in the order of Incubation, Centrifugation, Filtration, and then Aliquoting (if specified).
Default Calculation: Resolves to True if any of the corresponding Filter options are set. Otherwise, resolves to False.
FiltrationType
Default Calculation: Will automatically resolve to a filtration type appropriate for the volume of sample being filtered.
FilterInstrument
Default Calculation: Will automatically resolved to an instrument appropriate for the filtration type.
Pattern Description: An object of type or subtype Model[Instrument, FilterBlock], Object[Instrument, FilterBlock], Model[Instrument, PeristalticPump], Object[Instrument, PeristalticPump], Model[Instrument, VacuumPump], Object[Instrument, VacuumPump], Model[Instrument, Centrifuge], Object[Instrument, Centrifuge], Model[Instrument, SyringePump], or Object[Instrument, SyringePump] or Null.
Programmatic Pattern: (ObjectP[{Model[Instrument, FilterBlock], Object[Instrument, FilterBlock], Model[Instrument, PeristalticPump], Object[Instrument, PeristalticPump], Model[Instrument, VacuumPump], Object[Instrument, VacuumPump], Model[Instrument, Centrifuge], Object[Instrument, Centrifuge], Model[Instrument, SyringePump], Object[Instrument, SyringePump]}] | Automatic) | Null
Filter
The filter that should be used to remove impurities from the SamplesIn prior to starting the experiment.
Default Calculation: Will automatically resolve to a filter appropriate for the filtration type and instrument.
Pattern Description: An object of type or subtype Model[Container, Plate, Filter], Model[Container, Vessel, Filter], or Model[Item, Filter] or Null.
Programmatic Pattern: (ObjectP[{Model[Container, Plate, Filter], Model[Container, Vessel, Filter], Model[Item, Filter]}] | Automatic) | Null
FilterMaterial
The membrane material of the filter that should be used to remove impurities from the SamplesIn prior to starting the experiment.
Default Calculation: Resolves to an appropriate filter material for the given sample is Filtration is set to True.
Pattern Description: Cellulose, Cotton, Polyethylene, Polypropylene, PTFE, Nylon, PES, PLUS, PVDF, GlassFiber, GHP, UHMWPE, EPDM, DuraporePVDF, GxF, ZebaDesaltingResin, NickelResin, Silica, or HLB or Null.
PrefilterMaterial
The material from which the prefilter filtration membrane should be made of to remove impurities from the SamplesIn prior to starting the experiment.
Pattern Description: Cellulose, Cotton, Polyethylene, Polypropylene, PTFE, Nylon, PES, PLUS, PVDF, GlassFiber, GHP, UHMWPE, EPDM, DuraporePVDF, GxF, ZebaDesaltingResin, NickelResin, Silica, or HLB or Null.
FilterPoreSize
The pore size of the filter that should be used when removing impurities from the SamplesIn prior to starting the experiment.
Default Calculation: Resolves to an appropriate filter pore size for the given sample is Filtration is set to True.
Pattern Description: 0.008 micrometers, 0.1 micrometers, 0.2 micrometers, 0.22 micrometers, 0.45 micrometers, 1. micrometer, 1.1 micrometers, 2.5 micrometers, 6. micrometers, 20. micrometers, 30. micrometers, or 100. micrometers or Null.
PrefilterPoreSize
The pore size of the filter; all particles larger than this should be removed during the filtration.
Pattern Description: 0.008 micrometers, 0.1 micrometers, 0.2 micrometers, 0.22 micrometers, 0.45 micrometers, 1. micrometer, 1.1 micrometers, 2.5 micrometers, 6. micrometers, 20. micrometers, 30. micrometers, or 100. micrometers or Null.
FilterSyringe
Default Calculation: Resolves to an syringe appropriate to the volume of sample being filtered, if Filtration is set to True.
Pattern Description: An object of type or subtype Model[Container, Syringe] or Object[Container, Syringe] or a prepared sample or Null.
Programmatic Pattern: ((ObjectP[{Model[Container, Syringe], Object[Container, Syringe]}] | _String) | Automatic) | Null
FilterHousing
The filter housing that should be used to hold the filter membrane when filtration is performed using a standalone filter membrane.
Default Calculation: Resolve to an housing capable of holding the size of the membrane being used, if filter with Membrane FilterType is being used and Filtration is set to True.
Pattern Description: An object of type or subtype Model[Instrument, FilterHousing], Object[Instrument, FilterHousing], Model[Instrument, FilterBlock], or Object[Instrument, FilterBlock] or Null.
Programmatic Pattern: (ObjectP[{Model[Instrument, FilterHousing], Object[Instrument, FilterHousing], Model[Instrument, FilterBlock], Object[Instrument, FilterBlock]}] | Automatic) | Null
FilterIntensity
Default Calculation: Will automatically resolve to 2000 GravitationalAcceleration if FiltrationType is Centrifuge and Filtration is True.
Pattern Description: Greater than 0 revolutions per minute or greater than 0 standard accelerations due to gravity on the surface of the earth or Null.
Programmatic Pattern: ((GreaterP[0*RPM] | GreaterP[0*GravitationalAcceleration]) | Automatic) | Null
FilterTime
Default Calculation: Will automatically resolve to 5 Minute if FiltrationType is Centrifuge and Filtration is True.
FilterTemperature
The temperature at which the centrifuge chamber will be held while the samples are being centrifuged during filtration.
Default Calculation: Will automatically resolve to 22 Celsius if FiltrationType is Centrifuge and Filtration is True.
FilterContainerOut
The desired container filtered samples should be produced in or transferred into by the end of filtration, with indices indicating grouping of samples in the same plates, if desired.
Default Calculation: Automatically set as the PreferredContainer for the Volume of the sample. For plates, attempts to fill all wells of a single plate with the same model before using another one.
Pattern Description: An object of type or subtype Model[Container] or Object[Container] or a prepared sample or {Index, Container} or Null.
Programmatic Pattern: (((ObjectP[{Model[Container], Object[Container]}] | _String) | {GreaterEqualP[1, 1] | Automatic, (ObjectP[{Model[Container], Object[Container]}] | _String) | Automatic}) | Automatic) | Null
FilterAliquotDestinationWell
The desired position in the corresponding AliquotContainer in which the aliquot samples will be placed.
Default Calculation: Automatically resolves to A1 in containers with only one position. For plates, fills wells in the order provided by the function AllWells.
FilterAliquotContainer
The desired type of container that should be used to prepare and house the filter samples which should be used in lieu of the SamplesIn for the experiment.
Programmatic Pattern: ((ObjectP[Model[Container]] | {GreaterEqualP[1, 1] | (Automatic | Null), (ObjectP[{Model[Container], Object[Container]}] | _String) | Automatic}) | Automatic) | Null
FilterAliquot
The amount of each sample that should be transferred from the SamplesIn into the FilterAliquotContainer when performing an aliquot before filtration.
Default Calculation: Automatically set as the smaller between the current sample volume and the maximum volume of the destination container.
Pattern Description: All or greater than or equal to 1 microliter and less than or equal to 20 liters or Null.
FilterSterile
Default Calculation: Resolve to False if Filtration is indicated. If sterile filtration is desired, this option must manually be set to True.
Aliquoting
Aliquot
Indicates if aliquots should be taken from the SamplesIn and transferred into new AliquotSamples used in lieu of the SamplesIn for the experiment. Note that if NumberOfReplicates is specified this indicates that the input samples will also be aliquoted that number of times. Note that Aliquoting (if specified) occurs after any Sample Preparation (if specified).
AliquotAmount
Default Calculation: Automatically set as the smaller between the current sample volume and the maximum volume of the destination container if a liquid, or the current Mass or Count if a solid or counted item, respectively.
Programmatic Pattern: ((RangeP[1*Microliter, 20*Liter] | RangeP[1*Milligram, 20*Kilogram] | GreaterP[0*Unit, 1*Unit] | GreaterP[0., 1.] | All) | Automatic) | Null
TargetConcentration
The desired final concentration of analyte in the AliquotSamples after dilution of aliquots of SamplesIn with the ConcentratedBuffer and BufferDiluent which should be used in lieu of the SamplesIn for the experiment.
TargetConcentrationAnalyte
Default Calculation: Automatically set to the first value in the Analytes field of the input sample, or, if not populated, to the first analyte in the Composition field of the input sample, or if none exist, the first identity model of any kind in the Composition field.
Pattern Description: An object of type or subtype Model[Molecule], Model[Molecule, cDNA], Model[Molecule, Oligomer], Model[Molecule, Transcript], Model[Molecule, Protein], Model[Molecule, Protein, Antibody], Model[Molecule, Carbohydrate], Model[Molecule, Polymer], Model[Resin], Model[Resin, SolidPhaseSupport], Model[Lysate], Model[ProprietaryFormulation], Model[Virus], Model[Cell], Model[Cell, Mammalian], Model[Cell, Bacteria], Model[Cell, Yeast], Model[Tissue], Model[Material], or Model[Species] or Null.
AssayVolume
Default Calculation: Automatically determined based on Volume and TargetConcentration option values.
Pattern Description: Greater than or equal to 1 microliter and less than or equal to 20 liters or Null.
ConcentratedBuffer
The concentrated buffer which should be diluted by the BufferDilutionFactor in the final solution (i.e., the combination of the sample, ConcentratedBuffer, and BufferDiluent). The ConcentratedBuffer and BufferDiluent will be combined and then mixed with the sample, where the combined volume of these buffers is the difference between the AliquotAmount and the total AssayVolume.
Pattern Description: An object of type or subtype Model[Sample] or Object[Sample] or a prepared sample or Null.
BufferDilutionFactor
The dilution factor by which the concentrated buffer should be diluted in the final solution (i.e., the combination of the sample, ConcentratedBuffer, and BufferDiluent). The ConcentratedBuffer and BufferDiluent will be combined and then mixed with the sample, where the combined volume of these buffers is the difference between the AliquotAmount and the total AssayVolume.
Default Calculation: If ConcentratedBuffer is specified, automatically set to the ConcentratedBufferDilutionFactor of that sample; otherwise, set to Null.
BufferDiluent
The buffer used to dilute the aliquot sample such that ConcentratedBuffer is diluted by BufferDilutionFactor in the final solution. The ConcentratedBuffer and BufferDiluent will be combined and then mixed with the sample, where the combined volume of these buffers is the difference between the AliquotAmount and the total AssayVolume.
Default Calculation: Automatically resolves to Model[Sample, "Milli-Q water"] if ConcentratedBuffer is specified; otherwise, resolves to Null.
Pattern Description: An object of type or subtype Model[Sample] or Object[Sample] or a prepared sample or Null.
AssayBuffer
The buffer that should be added to any aliquots requiring dilution, where the volume of this buffer added is the difference between the AliquotAmount and the total AssayVolume.
Default Calculation: Automatically resolves to Model[Sample, "Milli-Q water"] if ConcentratedBuffer is not specified; otherwise, resolves to Null.
Pattern Description: An object of type or subtype Model[Sample] or Object[Sample] or a prepared sample or Null.
AliquotSampleStorageCondition
The non-default conditions under which any aliquot samples generated by this experiment should be stored after the protocol is completed.
Pattern Description: {AmbientStorage, Refrigerator, Freezer, DeepFreezer, CryogenicStorage, YeastIncubation, YeastShakingIncubation, BacterialIncubation, BacterialShakingIncubation, MammalianIncubation, ViralIncubation, CrystalIncubation, AcceleratedTesting, IntermediateTesting, LongTermTesting, UVVisLightTesting} or Disposal or Null.
DestinationWell
The desired position in the corresponding AliquotContainer in which the aliquot samples will be placed.
Default Calculation: Automatically resolves to A1 in containers with only one position. For plates, fills wells in the order provided by the function AllWells.
Pattern Description: Any well from A1 to H12 or list of one or more any well from A1 to H12 or any well from A1 to H12 entries or Null.
Programmatic Pattern: ((WellPositionP | {((Automatic | Null) | WellPositionP)..}) | Automatic) | Null
AliquotContainer
The desired type of container that should be used to prepare and house the aliquot samples, with indices indicating grouping of samples in the same plates, if desired. This option will resolve to be the length of the SamplesIn * NumberOfReplicates.
Default Calculation: Automatically set as the PreferredContainer for the AssayVolume of the sample. For plates, attempts to fill all wells of a single plate with the same model before aliquoting into the next.
Pattern Description: An object of type or subtype Model[Container] or Object[Container] or a prepared sample or Automatic or Null or {Index, Container} or list of one or more an object of type or subtype Model[Container] or Object[Container] or a prepared sample or Automatic or Null entries or list of one or more Automatic or Null or {Index, Container} entries.
Programmatic Pattern: (((ObjectP[{Model[Container], Object[Container]}] | _String) | (Automatic | Null) | {GreaterEqualP[1, 1] | (Automatic | Null), (ObjectP[{Model[Container], Object[Container]}] | _String) | (Automatic | Null)} | {((ObjectP[{Model[Container], Object[Container]}] | _String) | (Automatic | Null))..} | {({GreaterEqualP[1, 1] | (Automatic | Null), (ObjectP[{Model[Container], Object[Container]}] | _String) | (Automatic | Null)} | (Automatic | Null))..}) | Automatic) | Null
AliquotPreparation
Default Calculation: Automatic resolution will occur based on manipulation volumes and container types.
ConsolidateAliquots
Protocol Options
Organizational Information
Template
A template protocol whose methodology should be reproduced in running this experiment. Option values will be inherited from the template protocol, but can be individually overridden by directly specifying values for those options to this Experiment function.
Pattern Description: An object of type or subtype Object[Protocol] or an object of type or subtype of Object[Protocol] with UnresolvedOptions, ResolvedOptions specified or Null.
Programmatic Pattern: (ObjectP[Object[Protocol]] | FieldReferenceP[Object[Protocol], {UnresolvedOptions, ResolvedOptions}]) | Null
Name
A object name which should be used to refer to the output object in lieu of an automatically generated ID number.
Post Experiment
MeasureWeight
Indicates if any solid samples that are modified in the course of the experiment should have their weights measured and updated after running the experiment. Please note that public samples are weighed regardless of the value of this option.
MeasureVolume
Indicates if any liquid samples that are modified in the course of the experiment should have their volumes measured and updated after running the experiment. Please note that public samples are volume measured regardless of the value of this option.
ImageSample
Example Calls
Basics
High performance liquid chromatography (HPLC) separates sample mixtures into analyzable molecular constituents by injection into flowing liquid that passes through a retentive column:
Separation Mode and Gradient
Various modalities are available to elicit separation in HPLC including changing the composition of the buffers over time. For example, often ReversePhase is used to separate a mixture of polar and non-polar compounds. ReversePhase entails using a non-polar resin in the Column and linearly changing the mobile phase from polar to non-polar, solvating moieties adsorbed to the column. To run a ReversePhase separation, simply run:
Standards and Blanks
ExperimentHPLC can be used to quantify analytes in a sample, in which case, a Standard sample is employed to serve as reference. A standard can be submitted before and after the injection sequence of the samples simply by running:
Likewise, a Blank sample can be used to see if there is any background from the injection process. To run a blank to occur between every 5 samples, use the following command:
Fraction Collection
ExperimentHPLC can physically collect the separated analytes, based on the properties of the resulting detection signal. To turn this option on, run the following command:
Parameters can be set to control the triggering of FractionCollection. Here fractions are collected from peaks that exceed a minimum absorbance value:
FractionCollectionDetector can be set to trigger fraction collection based on the peaks in the desired channel:
PhotoDiodeArray detection
PhotoDiodeArray detection involves passing light across a range of wavelengths and measuring how much light is absorbed by the sample at each even wavelength. To run a PhotoDiodeArray HPLC experiment, simply run:
Fluorescence detection
Fluorescence detection involves the excitation of the mobile phase with light at defined wavelengths. Fluorescent moieties within the mobile phase will accordingly emit light, also measurable at defined wavelengths. Therefore, directly defining excitation and emission wavelengths is possible for successful measurement. To run a fluorescence HPLC experiment, simply run:
Evaporative Light Scattering detection
Evaporative light scattering detection (ELSD) measurement forms dispersed droplets from the column effluent. The droplets are formed through nebulization, which is accomplished through a combination of heat and a sheath gas. Droplets containing analytes are different sizes, and this variability in droplet size is measurable through light scattering. The sheath gas carries the droplets through the heated drift tube for eventual scattering measurement. Accordingly, the ELSD measurement directly measures this light scattering over time of the measurement. An ELSD experiment can be executed by simply running:
ELSD provides detailed control over how the scattering functions, and the flow rate of the sheath gas and the temperature of the drift tube can be specified. For example, to increase the flow rate and control the temperature, the options may be specified as follows:
pH and conductance detection
pH and conductance detectors monitor the pH and conductance values of the flow in the HPLC system to verify elution conditions, sample retention behavior, and instrument precision during analysis:
pH and conductance measurements can be automatically corrected according to the temperatures inside the corresponding flow cells:
Multi-angle light scattering (MALS) and dynamic light scattering (DLS) detection
In the Multi-angle light scattering (MALS) and dynamic light scattering (DLS) detector, laser light is scattered by the nanoparticles or biomacromolecules in the sample inside the flow cell and the intensities and fluctuation of the scattered light beams at different directions are measured. The results from MALS and DLS detection can provide analysis of molar mass, radius of gyration and hydrodynamic radius for copolymers and protein conjugates:
Refractive index (RI) detection
Refractive index detection involves the measurement of the shift of the light beam direction across the sample flow:
RefractiveIndexMethod can be set to DifferentialRefractiveIndex to measure the refractive index difference between the sample and the reference (loaded in the previous gradient step) in order to determine the concentration of the sample:
Injection Table
Preferred Input Containers
The Watars H-Class instrument, Ultimate 3000 instrument, and Agilent 1260 Infinity II Semi-Preparative HPLC instrument autosamplers can take 2mL deep well plates.
The Watars H-Class instrument, Ultimate 3000 instrument, and Agilent 1260 Infinity II Semi-Preparative HPLC instrument autosamplers can take normal vials routinely used in HPLC analysis.
Data Processing
Warnings and Errors
Messages (160)
AbsorbanceRateAdjusted (1)
BlankFrequencyNoBlanks (1)
BlankOptionsButNoFrequency (1)
BufferDMustExistForGradient (1)
BufferDMustExistForInstrument (1)
BufferDMustExistForSampleTemperature (1)
ColumnGap (2)
ColumnOptionsButNoColumn (1)
ColumnOrientationSelector (1)
ColumnPositionColumnConflict (1)
ColumnPositionInjectionTableConflict (1)
ColumnSelectorConflict (1)
ColumnSelectorInstrumentConflict (1)
ColumnTemperatureInjectionTableConflict (1)
ConflictColumnStorageBuffer (1)
ConflictColumnStorageBufferFlushGradient (1)
ConflictFractionCollectionUnit (1)
ConflictFractionOptionSpecification (1)
ConflictHPLCDetectorOptions (1)
ConflictHPLCFluorescenceOptionsLengths (1)
ConflictingFractionCollectionMethodOptions (1)
ConflictingInjectionSampleVolumeMeasurementOption (1)
ConflictRefractiveIndexMethod (1)
ConflictScaleAndCollectFractions (1)
DetectorConflict (2)
DuplicateColumnSelectorPositions (1)
FractionCollectionWavelengthConflict (1)
GasHeatingRequiresNebulizer (1)
GasPressureRequiresNebulizer (1)
HeatingPowerRequiresNebulizerHeating (1)
HPLCBufferConflict (1)
HPLCCannotIncubateColumn (1)
HPLCColumnsCannotFit (1)
HPLCConflictingFractionCollectionOptions (1)
HPLCEmissionExcitationTooNarrow (3)
If Dionex Fluorescence HPLC is required, excitation wavelength should not be within 20 nm of emission wavelength:
If semi-prep Agilent Fluorescence HPLC is required, excitation wavelength must be 10 nm larger than emission wavelength (2):
If semi-prep Agilent Fluorescence HPLC is required, excitation wavelength should not be within 10 nm of emission wavelength (1):
HPLCEmissionLowerThanExcitation (1)
HPLCFluorescenceWavelengthLimit (2)
HPLCGradientNotReequilibrated (1)
HPLCIncompatibleAliquotContainer (1)
HPLCIncompatibleInjectionVolume (2)
HPLCInstrumentScaleConflict (1)
HPLCpHCalibrationBufferSwapped (1)
HPLCSmallInjectionVolume (1)
HPLCTooManySamples (1)
IncompatibleBlankFlowRate (1)
IncompatibleColumnFlushFlowRate (1)
IncompatibleColumnPrimeFlowRate (1)
IncompatibleColumnTechnique (2)
IncompatibleColumnTemperature (2)
IncompatibleColumnType (3)
If multiple columns are specified, a warning is thrown if any specified Column's SeparationMode does not match the specified Type:
Warning is thrown if specified Column's SeparationMode does not match the specified SeparationMode:
Warning is thrown if specified GuardColumn's SeparationMode does not match the specified SeparationMode:
IncompatibleContainerAndNeedleWashBuffer (1)
IncompatibleContainerModel (1)
IncompatibleDetectionWavelength (3)
Error if the specified Fluorescence Detector is not compatible with any instrument that supports the emission wavelength(s) specified:
Error if the specified Fluorescence Detector is not compatible with any instrument that supports the excitation wavelength(s) specified:
Error if the specified UVVis Detector is not compatible with any instrument that supports the absorbance detection wavelength(s) specified:
IncompatibleFlowRate (1)
IncompatibleFractionCollectionAndNeedleWashBuffer (1)
IncompatibleHPLCColumnTemperature (3)
Error if the selected instrument is not available to meet the specified column temperature from the specified gradient methods (or no instrument can meet the requirements):
Error if the selected instrument is not available to meet the specified column temperature inside the column temperature options (or no instrument can meet the requirements):
Error if the selected instrument is not available to meet the specified column temperature inside the InjectionTable (or no instrument can meet the requirements):
IncompatibleInstrumentBufferD (1)
IncompatibleModelColumnStorageBuffer (1)
IncompatibleNeedleWash (1)
IncompatibleStandardFlowRate (1)
InjectionTableBlankFrequencyConflict (1)
InjectionTableColumnConflictHPLC (1)
InjectionTableForeignSamples (1)
InjectionTableGradientConflict (1)
InjectionTableStandardConflict (1)
InjectionTableStandardFrequencyConflict (1)
InjectionVolumeConflict (1)
InstrumentPrecision (8)
Warning will be thrown if a buffer composition is specified to an incompatible precision:
Warning will be thrown if a gradient flow rate is specified to an incompatible precision:
Warning will be thrown if a gradient flow rate is specified to an incompatible precision:
Warning will be thrown if a gradient timepoint is specified to an incompatible precision:
Warning will be thrown if a injection volume is specified to an incompatible precision:
Warning will be thrown if an absorbance is specified to an incompatible precision:
Warning will be thrown if an absorbance slope is specified to an incompatible precision:
Warning will be thrown if a temperature is specified to an incompatible precision:
InsufficientSampleVolume (2)
InvalidFractionCollectionContainer (2)
InvalidFractionCollectionEndTime (1)
InvalidGradientCompositionOptions (1)
InvalidHPLCAlternateInstruments (2)
InvalidHPLCConductivityCalibrationOptions (1)
InvalidHPLCDetectorOptions (1)
InvalidHPLCEmissionCutOffFilter (1)
InvalidHPLCFluorescenceFlowCellTemperature (1)
InvalidHPLCMaxAcceleration (1)
InvalidHPLCpHCalibrationOptions (1)
InvalidWatersHPLCFluorescenceGain (1)
MissingFractionCollectionDetector (1)
MissingHPLCConductivityCalibrationOptions (2)
MissingHPLCDetectorOptions (1)
MissingHPLCpHCalibrationOptions (2)
MissingHPLCScaleInstrument (1)
NoSuitableInstrumentForDetection (1)
NotApplicableHPLCPeakFractionCollectionOptions (1)
ObjectDoesNotExist (4)
OverwritingGradient (1)
RemovedExtraGradientEntries (1)
SamplesOutStorageConditionRequired (1)
SampleTemperatureConflict (1)
StandardFrequencyNoStandards (1)
StandardOptionsButNoFrequency (1)
StandardOptionsButNoStandard (1)
StandardsButNoFrequency (1)
TooLargeHPLCEmissionCutOffFilter (1)
UnsupportedBufferD (1)
UnsupportedBufferDAndDetectors (1)
UnsupportedGradientD (1)
UnsupportedGradientDAndDetectors (1)
UnsupportedSampleTemperature (1)
UnsupportedSampleTemperatureAndDetectors (1)
UVVisOptionsNotApplicable (1)
VariableColumnTypes (1)
WavelengthOutOfRange (3)
Error if a detection wavelength is specified that is outside the specified Instrument's compatible range:
Error if an excitation wavelength is specified that is outside the specified Instrument's compatible range:
Error if an excitation wavelength is specified that is outside the specified Instrument's compatible range:
WavelengthResolutionAdjusted (1)
WavelengthResolutionConflict (1)
Possible Issues
Equilibration
Equilibrate to the correct buffer conditions at the end of each gradient to reach a state that is appropriate for the start of the next gradient.
Column flush
The column will end up being stored in whatever the final gradient solvent conditions are so that should be considered for column protection.
Column fouling
Consider using a GuardColumn to preserve column lifetime. Examine injections of standards over time to ensure that the column is performing to satisfaction.
Filter sample and buffer
Last modified on Sun 23 Feb 2025 21:51:41