Nephelometry
Nephelometry[Options]⟹UnitOperation
measures the nephelometry data of the input samples.
Nephelometry is a measurement of the intensity of scattered light upon passing through a solution containing insoluble, suspended particles. Samples are exposed to visible light at a specific wavelength, and the intensity of the light that is scattered by the particles in solution is measured. The intensity of scattered light is correlated to the amount of particulate matter suspended in solution by comparison to a standard curve. Nephelometry is often used for immunology applications, measuring amounts of antigen in a sample by adding known amounts of antibody, measuring purity of water, drug solubility screening, and measuring turbidity of cell cultures.
Experimental Principles
Figure 1.1: Procedural overview of an Nephelometry experiment using the NEPHELOstar Plus plate reader. Step 1: Plate is loaded with sample. Step 2: The plate is inserted into the plate reader. Step 3: Injection samples are injected into the plate if specified. Step 4: The turbidity measurement is collected for each of the samples.
Instrumentation
NEPHELOstar Plus
Figure 2.1: Instrument schematic of a BMG NEPHELOstar Plus nephelometer plate reader. In the NEPHELOstar Plus, a laser sends photons through each well of the plate. The light that is scattered from particles in solution is then directed through a Ulbricht sphere which collects the scattered light and directs it to be measured by a photodiode detector.
The plate chamber can be heated up to 65°C and it can mix the plate at up to 700 RPM before the run.
The reader has two 300μL syringe pump injectors which can be used for 0.5 - 300 μL injections of two unique samples prior to the run.
Experiment Options
General
Sample
Programmatic Pattern: ((ObjectP[{Object[Sample], Object[Container]}] | _String) | {Alternatives @@ Flatten[AllWells[NumberOfWells -> 384]] | LocationPositionP, ObjectP[{Object[Container]}] | _String} | ObjectP[Model[Sample]]) | Null
SampleLabel
A user defined word or phrase used to identify the samples for which nephelometry measurements are taken, for use in downstream unit operations.
SampleContainerLabel
A user defined word or phrase used to identify the containers of the samples for which nephelometry measurements are taken, for use in downstream unit operations.
Analyte
If the Method->Solublity, the compound whose concentration should be determined during this experiment. If Method->CellCount, the strain of cells whose concentration will be determined during this experiment, based on a previously measured standard curve, ReferenceCellCountCurve in the Model[Cell] composing the sample.
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.
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]
BlankLabel
A user defined word or phrase used to identify the blank samples, for use in downstream unit operations.
Instrument
Pattern Description: An object of type or subtype Model[Instrument, Nephelometer] or Object[Instrument, Nephelometer]
Method
The type of experiment nephelometric measurements are collected for. CellCount involves measuring the amount of scattered light from cells suspended in solution and using a Standard Curve that relates RNU (Relative Nephelometric Units) to Cells/mL in order to quantify the number of cells in solution. For Method -> Solubility, scattered light from suspended particles in solution will be measured, at a single time point or over a time period. Reagents can be injected into the samples to study their effects on solubility, and dilutions can be performed to determine the point of saturation.
Default Calculation: Automatically set to CellCount if samples are cells and have a StandardCurve. Otherwise, Method is set to Solubility.
PreparedPlate
Indicates if a prepared plate is provided for nephelometric measurement. The prepared plate contains the solutions that are ready for tubidity measurements in a plate reader. If 'PreparedPlate' is True, dilution options and Moat options must be Null.
RetainCover
Indicates if the plate seal or lid on the assay container should not be taken off during measurement to decrease evaporation. When this option is set to True, injections cannot be performed as it's not possible to inject samples through the cover. The cover must be clear, as the laser passes from the top of the plate through to the bottom of the plate.
Preparation
Indicates if this unit operation is carried out primarily robotically or manually. Manual unit operations are executed by a laboratory operator and robotic unit operations are executed by a liquid handling work cell.
WorkCell
The automated workstation with a collection of integrated instruments on which this unit operation will be will be performed if Preparation -> Robotic.
NumberOfReplicates
The number of times to repeat nephelometry reading on each provided sample. If Aliquot -> True, this also indicates the number of times each provided sample will be aliquoted.
Quantification
QuantifyCellCount
Indicates if the number of cells in the sample is automatically converted from the measured RNU (Relative Nephelometric Unit) via a StandardCurve in the Model[Cell].
Default Calculation: Automatically set to True if Method->CellCount, and set to False if Method->Solubility.
InputConcentration
Default Calculation: Automatically determined from the concentration of the analyte in the original source sample.
Sample Preparation
DilutionCurve
The collection of dilutions that will be performed on the samples before nephelometric measurements are taken. For Fixed Dilution Volume Dilution Curves, the Sample Amount is the volume of the sample that will be mixed with the Diluent Volume of the Diluent to create the desired concentration. For Fixed Dilution Factor Dilution Curves, the Sample Volume is the volume of the sample that will created after being diluted by the Dilution Factor. For example, a 1M sample with a dilution factor of 0.7 will be diluted to a concentration 0.7M. IMPORTANT: Because the dilution curve does not intrinsically include the original sample, in the case of sample dilution the first diluting factor should be 1 or Diluent Volume should be 0 Microliter to include the original sample. If dilutions and injections are specified, injection samples will be injected into every dilution in the curve corresponding to SamplesIn.
Figure 3.1: Use the DilutionCuve option to create a collection of dilutions that will be performed on the sample.
Default Calculation: This is automatically set Null if Diluent is set to Null or a Serial Dilution Curve is specified.
Programmatic Pattern: ((Null | ({{RangeP[0*Microliter, 300*Microliter], RangeP[0*Microliter, 300*Microliter]}..} | {{RangeP[0*Microliter, 300*Microliter], RangeP[0, 1]}..})) | Automatic) | Null
SerialDilutionCurve
The collection of dilutions that will be performed on the samples before nephelometric measurements are taken. For Serial Dilution Volumes, the Transfer Volume is taken out of the sample and added to a second well with the Diluent Volume of the Diluent. It is mixed, then the Transfer Volume is taken out of that well to be added to a third well. This is repeated to make Number Of Dilutions diluted samples. For example, if a 100 ug/ ml sample with a Transfer Volume of 20 Microliters, a Diluent Volume of 60 Microliters and a Number of Dilutions of 3 is used, it will create a DilutionCurve of 25 ug/ ml, 6.25 ug/ ml, and 1.5625 ug/ ml with each dilution having a volume of 60 Microliters. For Serial Dilution Factors, the sample will be diluted by the dilution factor at each transfer step. IMPORTANT: Because the dilution curve does not intrinsically include the original sample, in the case of sample dilution the first diluting factor should be 1 or Diluent Volume should be 0 Microliter to include the original sample. If dilutions and injections are specified, injection samples will be injected into every dilution in the curve corresponding to SamplesIn.
Figure 3.2: Use the SerialDilutionCuve option to create a collection of serial dilutions that will be performed on the sample.
Default Calculation: This is automatically set to Null if Diluent is set to Null or a non-serial Dilution Curve is specified. In all other cases it is automatically set to TransferVolume as one tenth of smallest of sample volume or container max volume, DiluentVolume as smallest of sample volume or container max volume, and Number of Dilutions as 3.
Programmatic Pattern: ((Null | ({RangeP[0*Microliter, 300*Microliter], RangeP[0*Microliter, 300*Microliter], GreaterP[1, 1]} | {RangeP[0*Microliter, 300*Microliter], {RangeP[0, 1], GreaterP[1, 1]} | {RangeP[0, 1]..}})) | Automatic) | Null
Diluent
Default Calculation: If the measurement is to be performed with DilutionCurve Null, this is automatically set to Null. Otherwise, it is automatically set to the Solvent of the sample. If the Solvent field is not informed, Diluent is set to Model[Sample,"Milli-Q water"].
Pattern Description: An object of type or subtype Model[Sample] or Object[Sample] or a prepared sample or Null.
Programmatic Pattern: ((Null | (ObjectP[{Model[Sample], Object[Sample]}] | _String)) | Automatic) | Null
Injections
PrimaryInjectionSample
The sample to be injected in the first round of injections in order to introduce a time sensitive reagent/sample into the plate before/during measurement. The corresponding injection volumes can be set with PrimaryInjectionVolume. If DilutionCurve or SerialDilutionCurve is specified, PrimaryInjectionSample will be injected into all dilutions in the curve.
Pattern Description: An object of type or subtype Object[Sample] or Model[Sample] or a prepared sample or Null.
PrimaryInjectionVolume
Pattern Description: Greater than or equal to 0.5 microliters and less than or equal to 300 microliters or Null.
SecondaryInjectionSample
The sample to be injected in the second round of injections. Set the corresponding injection volumes with SecondaryInjectionVolume. If DilutionCurve or SerialDilutionCurve is specified, SecondaryInjectionSample will be injected into all dilutions in the curve.
Pattern Description: An object of type or subtype Object[Sample] or Model[Sample] or a prepared sample or Null.
SecondaryInjectionVolume
Pattern Description: Greater than or equal to 0.5 microliters and less than or equal to 300 microliters or Null.
PrimaryInjectionFlowRate
The speed at which to transfer injection samples into the assay plate in the first round of injections.
Pattern Description: 430 microliters per second, 400 microliters per second, 350 microliters per second, 300 microliters per second, 260 microliters per second, 220 microliters per second, 190 microliters per second, 170 microliters per second, 150 microliters per second, 135 microliters per second, 115 microliters per second, 100 microliters per second, 85 microliters per second, 65 microliters per second, or 50 microliters per second or Null.
SecondaryInjectionFlowRate
The speed at which to transfer injection samples into the assay plate in the second round of injections.
Pattern Description: 430 microliters per second, 400 microliters per second, 350 microliters per second, 300 microliters per second, 260 microliters per second, 220 microliters per second, 190 microliters per second, 170 microliters per second, 150 microliters per second, 135 microliters per second, 115 microliters per second, 100 microliters per second, 85 microliters per second, 65 microliters per second, or 50 microliters per second or Null.
PrimaryInjectionSampleStorageCondition
The non-default conditions under which any injection samples used by this experiment are stored after the protocol is completed.
Pattern Description: {AmbientStorage, EnclosedAmbientStorage, Refrigerator, Freezer, DeepFreezer, CryogenicStorage, YeastIncubation, YeastShakingIncubation, BacterialIncubation, BacterialShakingIncubation, MammalianIncubation, ViralIncubation, CrystalIncubation, AcceleratedTesting, IntermediateTesting, LongTermTesting, UVVisLightTesting} or Disposal or Null.
SecondaryInjectionSampleStorageCondition
The non-default conditions under which any injection samples used by this experiment are stored after the protocol is completed.
Pattern Description: {AmbientStorage, EnclosedAmbientStorage, Refrigerator, Freezer, DeepFreezer, CryogenicStorage, YeastIncubation, YeastShakingIncubation, BacterialIncubation, BacterialShakingIncubation, MammalianIncubation, ViralIncubation, CrystalIncubation, AcceleratedTesting, IntermediateTesting, LongTermTesting, UVVisLightTesting} or Disposal or Null.
Data Processing
Blank
The source used to generate a blank sample whose scattered light is subtracted as background from the scattered light readings of the input sample.
Default Calculation: Automatically set to Null if BlankMeasurement is False. If BlankMeasurement->True, Blank is set to the Solvent field for each sample. If the Solvent field is not informed, Blank is set to Model[Sample,"Milli-Q water"].
Pattern Description: An object of type or subtype Model[Sample] or Object[Sample] or a prepared sample or Null.
Programmatic Pattern: ((Null | (ObjectP[{Model[Sample], Object[Sample]}] | _String)) | Automatic) | Null
BlankVolume
The volume of the blank that should be transferred out and used for blank measurements. Set BlankVolume to Null to indicate blanks should be read inside their current containers.
Default Calculation: If BlankMeasurement is True, automatically set to match the maximum of the volume of each sample or the recommended fill volume of the container. Otherwise set to Null.
Pattern Description: Greater than or equal to 1 microliter and less than or equal to 4000 microliters or Null.
BlankMeasurement
Indicates if blank samples are prepared to account for the background signal when measuring scattered light from the assay samples.
Post Experiment
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, EnclosedAmbientStorage, Refrigerator, Freezer, DeepFreezer, CryogenicStorage, YeastIncubation, YeastShakingIncubation, BacterialIncubation, BacterialShakingIncubation, MammalianIncubation, ViralIncubation, CrystalIncubation, AcceleratedTesting, IntermediateTesting, LongTermTesting, UVVisLightTesting} or Disposal or Null.
Optics
BeamAperture
The diameter of the opening allowing the source laser light to pass through to the sample. A larger BeamAperture allows more light to pass through to the sample, leading to a higher signal. A setting of 1.5 millimeters is recommended for all 384 and 96 well plates, and 2.5-3.5 millimeters for 48 or less well plates. For non-homogenous solutions, a higher BeamAperture is recommended, and for samples with a large meniscus effect, a smaller BeamAperture is recommended.
Pattern Description: Greater than or equal to 1.5 millimeters and less than or equal to 3.5 millimeters.
BeamIntensity
The percentage of the total amount of the laser source light passed through to reach the sample. For Solubility experiments, 80% is recommended, and for experiments with highly concentrated or highly turbid samples, such as those involving cells, a BeamIntensity of 10% is recommended.
Measurement
Temperature
Default Calculation: Automatically set to Ambient if Method->Solubility or the average of the IncubationTemperatures of the Model[Cell] Analytes, or 37 Celsius if that field is not informed if Method->CellCount.
Pattern Description: Ambient or greater than or equal to 25 degrees Celsius and less than or equal to 65 degrees Celsius.
EquilibrationTime
The length of time for which the assay plates equilibrate at the requested temperature in the plate reader before being read.
Default Calculation: Automatically set to 0 second when Temperature is set to Ambient, or 5 minutes when Temperature is above Ambient.
TargetCarbonDioxideLevel
Pattern Description: Greater than or equal to 0.1 percent and less than or equal to 20 percent or Null.
TargetOxygenLevel
The target amount of oxygen in the atmosphere in the plate reader chamber. If specified, nitrogen gas is pumped into the chamber to force oxygen in ambient air out of the chamber until the desired level is reached.
Pattern Description: Greater than or equal to 0.1 percent and less than or equal to 20 percent or Null.
PlateReaderMix
PlateReaderMixTime
The amount of time samples should be mixed inside the plate reader chamber before the samples are read.
Default Calculation: Automatically set to 30 second if PlateReaderMix is True or any other plate reader mix options are specified.
PlateReaderMixRate
The rate at which the plate is agitated inside the plate reader chamber before the samples are read.
Default Calculation: Automatically set to 700 RPM if PlateReaderMix is True or any other plate reader mix options are specified.
Pattern Description: Greater than or equal to 100 revolutions per minute and less than or equal to 700 revolutions per minute or Null.
PlateReaderMixMode
Default Calculation: Automatically set to DoubleOrbital if PlateReaderMix is True or any other plate reader mix options are specified.
MoatSize
Indicates the number of concentric perimeters of wells which should be filled with MoatBuffer in order to decrease evaporation from the assay samples during the run.
Figure 3.3: Use the moat options, MoatBuffer, MoatVolume and MoatSize to create an outer ring of wells filled with buffer. This has been shown to decrease evaporation during long reads.
Pattern Description: Greater than or equal to 1 and less than or equal to 7 in increments of 1 or Null.
MoatVolume
Default Calculation: Automatically set to the RecommendedFillVolume of the assay plate model, or if Null, then 75% of the maximum volume of the assay plate if any other moat options are specified.
MoatBuffer
Default Calculation: Automatically set to Model[Sample,"Milli-Q water"] if any other moat options are specified.
Pattern Description: An object of type or subtype Object[Sample] or Model[Sample] or a prepared sample or Null.
ReadDirection
Indicate the plate path the instrument will follow as it measures scattered light in each well, for instance reading all wells in a row before continuing on to the next row (Row).
Figure 3.4: Use the ReadDirection option to control how the plate reader scans the plate during each read cycle. Choosing a more efficient path will lead to a shorter minimum CycleTime.
SettlingTime
The time between the movement of the plate and the beginning of the measurement. It reduces potential vibration of the samples in plate due to the stop and go motion of the plate carrier.
ReadStartTime
The time at which nephelometry measurement readings will begin, after the plate is in position and the SettlingTime has passed. If a pause is desired before readings begin, set this time to higher than the SettlingTime. For kinetic experiments, the ReadStartTime is the time at which the first cycle begins.
IntegrationTime
The amount of time that scattered light is measured. Increasing the IntegrationTime leads to higher measurements.
Default Calculation: Automatically set to 1 second if no SamplingPattern is specified, or if SamplingPattern->Matrix. If SamplingPattern->Ring or Spiral, IntegrationTime is set based on the SamplingDistance.
Sample Handling
AtmosphereEquilibrationTime
The length of time for which the samples equilibrate at the requested oxygen and carbon dioxide level before being read.
Default Calculation: Automatically set to 5 Minute if TargetCarbonDioxideLevel or TargetOxygenLevel is specified. Otherwise, set to Null.
Sampling
SamplingDistance
Default Calculation: Automatically set to 80% of the well diameter of the assay container if SamplingPattern is not Null.
Pattern Description: Greater than or equal to 1 millimeter and less than or equal to 6 millimeters or Null.
SamplingPattern
Indicates where in the well measurements are taken. Center indicates that measurements are taken from the center of the well. Ring indicates a ring within the well with a outer diameter of SamplingDistance. Spiral indicates a spiral from the diameter of SamplingDistance to the center of the well. Matrix indicates a grid of readings matching SamplingDimension, filling SamplingDistance. If any part of the grid falls outside of the well, it will not be included in the measurements. Additionally, Spiral will return a much higher background and should only be used bacteria and yeast with a high propensity to clump.
Default Calculation: Resolves to Ring if SamplingDistance is set, and resolves to Matrix if SamplingDimension is set, otherwise resolves to Center.
SamplingDimension
Specifies the size of the grid used for Matrix sampling to fill to SamplingDistance. For example, SamplingDimension->5 will scan a 5 x 5 grid.
Sample Prep Options
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: ((Null | (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.
Programmatic Pattern: ((Null | ObjectP[Join[MixInstrumentModels, MixInstrumentObjects]]) | Automatic) | 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: ((Null | (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: ((Null | 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: ((Null | (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: ((Null | (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: ((Null | 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: ((Null | 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, AgaroseResin, CobaltResin, Silica, HLB, or AnoporeAlumina 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, AgaroseResin, CobaltResin, Silica, HLB, or AnoporeAlumina 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, 0.02 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, 0.02 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: ((Null | (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: ((Null | 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: ((Null | (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: ((Null | ((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: ((Null | (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).
AliquotSampleLabel
A user defined word or phrase used to identify the samples after they are aliquotted, for use in downstream unit operations.
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: ((Null | (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.
Programmatic Pattern: ((Null | (ObjectP[{Model[Sample], Object[Sample]}] | _String)) | Automatic) | 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.
Programmatic Pattern: ((Null | (ObjectP[{Model[Sample], Object[Sample]}] | _String)) | Automatic) | 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.
Programmatic Pattern: ((Null | (ObjectP[{Model[Sample], Object[Sample]}] | _String)) | Automatic) | 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, EnclosedAmbientStorage, 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
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
Method
Setting Method->Solubility will obtain a RNU (Relative Nephelometric Unit) measurement from the sample:
Cell samples that already have a Standard Curve (in the StandardCurves field of the Model[Cell]) that relates RNU to Cell/mL should use Method->CellCount to have the RNU value automatically be converted to Cell/mL. To obtain a Standard Curve, please call ExperimentQuantifyCells:
PreparedPlate
If samples and blanks are already in a plate on the list BMGCompatiblePlate[Nephelometry], and no moat or dilutions are desired, set PreparedPlate to True:
Analyte
Optics
Quantification
If QuantifyCellCount is set to True, each sample must have a Model[Cell] Analyte with a StandardCurve that can convert RNU to Cell/mL. The number of cells in each sample will be calculated based on the nephelometry reading and the previously determined fit in StandardCurves. If there are multiple standard curves that can convert between RNU and Cell/mL, the standard curve with the acquisition parameters (BeamAperture, BeamIntensity, IntegrationTime) closest to the acquisition parameters specified in the current experiment is used :
Set the sample to set as the blank to subtract when quantifying. If not specified, will be set to the Solvent of the sample, or water if Null:
Dilutions
To specify dilutions, use the DilutionCurve and SerialDilutionCurve options, depending on the type of dilution desired. Dilutions can be specified based on volumes, specifying the sample and diluent volumes for standard dilutions or the volume of the sample that should be transferred, the volume of diluent the sample should be added to and the number of times the sample should be diluted for serial dilutions:
Dilutions can also be specified based on dilution factors, either fixed or variable for a serial dilution curve. For standard dilutions, specify the volume of the sample and dilution factor at each point of the curve. For serial dilutions diluting by a fixed factor at each step, provide the volume of the sample, a dilution factor and the number of times the sample should be diluted. For serial dilutions diluting by variable factors at each step, provide the volume of the sample and a series of dilution factors.
Atmosphere
To regulate the atmosphere inside the sample chamber, use option TargetCarbonDioxideLevel and TargetOxygenLevel:
Plate Reader Mixing
Moat
A moat may be specified to limit evaporation during long read times; use the MoatSize, MoatVolume, and MoatBuffer options to specify:
Injections
To specify injections, use the InjectionSample and InjectionVolume options. The sample and volume options are index matched to the input samples allowing you to control the wells which receive injections and the volumes of those injections. In the example below, 50 Microliter of myInjectionSample1 is injected into each well, and then myInjectionSample2 is injected at different volumes into the first two wells but not the third:
Warnings and Errors
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Messages
Possible Issues
Impurities in plates
If a plate is scratched on the bottom, this can effect the light scattering reading. It is recommended to measure at least 4 replicates in case of effects from the plate.
Meniscus effects
The meniscus of a sample can change the light scattering intensity from a sample. Sample wells should be filled as close to the MaxVolume of the wells as possible to minimize meniscus effects. When comparing Blank data to sample data, if the sample contains detergent or other similar compounds that effect the meniscus, blank measurements can be higher than sample measurements, leading to negative measurements when the blank measurement is subtracted. In these cases, raw measurements with blank data not subtracted should be used.
Last modified on Thu 11 Sep 2025 10:34:24