1.1.1.1 Description
The Assay file describes the protocol parameters and factors (e.g. temperature, media solvent, concentration) associated with each assay and provide references to assay results including measurements, derived data files, and other file types. There can be multiple Assay files per study.
1.1.1.2 File Format
The ISA-TAB file format leverages a horizontal based spreadsheet format with column headers and row values. An example subset of this format is provided below. Please note, concepts specified within the bracket “[]” are variable depending on the type of assay.
<ac:structured-macro ac:name="unmigrated-wiki-markup" ac:schema-version="1" ac:macro-id="cec2e4ae-5648-417e-ba32-3a41ebe05301"><ac:plain-text-body><![CDATA[ |
Material Sample Name |
Assay Name |
Factor Value [temperature] |
Unit |
Measurement Value [size] |
Unit |
]]></ac:plain-text-body></ac:structured-macro> |
NCL-23-F1 |
Size by DLS |
25 |
Celsius |
7.4 |
nm |
||
NCL-23-F1 |
Size by DLS |
37 |
Celsius |
9.8 |
nm |
1.1.1.3 nano-TAB Extensions
In support of nano-TAB, the following extensions and constraints were applied to the ISA-TAB Assay file and are further described in Section 4.
Field |
Change |
Description |
Field Status |
||
<ac:structured-macro ac:name="unmigrated-wiki-markup" ac:schema-version="1" ac:macro-id="3637eb34-c3ea-4caa-8880-7d5cf796c587"><ac:plain-text-body><![CDATA[ |
Measurement Value[] |
Addition |
The Measurement Value is leveraged to capture derived data values associated with each assay. In ISA-TAB this is typically represented in a separate data file; however, in assays in which there are only a few measurements (e.g. Z-Average Size) and it is necessary to display the factors along with the measurements (e.g. temperature), the Measurement Value is leveraged. |
Optional |
|
Statistic |
Addition |
The ISA-TAB format allows for the specification of units (e.g. mg/mL) but not a statistic (e.g. mean, SD, etc.). |
Optional |
||
<ac:structured-macro ac:name="unmigrated-wiki-markup" ac:schema-version="1" ac:macro-id="e54c945b-f4b7-4263-8caf-5968a41be342"><ac:plain-text-body><![CDATA[ |
Factor Value[] |
Use NPO |
The NPO should be leveraged to obtain a list of valid Factor values which are typically stressors or experimental conditions such as temperature, media solvent, time, and concentration. |
Optional |
]]></ac:plain-text-body></ac:structured-macro> |
1.1 Assay file
The Assay file is a horizontally oriented spreadsheet. The file supports the following types of information:
s the sample name
s the assay protocol reference and protocol parameter values
s the assay name, performer, and date the assay was performed
s assay factor values including units and statistics if applicable
s assay measurement values including units and statistics if applicable
s derived data files or image files if applicable
Specific concepts often leveraged in the Assay file are provided below.
Sample Name - The unique identification name of the sample. The sample name can be a biospecimen, nanomaterial, small molecule, or other material.
Assay Protocol Ref - The name of the protocol used to perform the experiment.
Parameter Value [] -- Values of concepts associated with an experiment protocol that are not varied in the experiment. Parameters concepts are variable and described in brackets.
Performer - The full name of the individual performing the assay.
Date - The calendar day on which the assay was performed in the format YYYY-MM-DD.
Assay Name - The name of the assay performed.
Factor Value [] - An independent variable manipulated by the experimentalist with the intention to affect biological systems (i.e. stressor). Factor concepts are variable and described in brackets.
Measurement Value [] - The endpoint of the assay. Measurement value concepts are variable and described in brackets.
Unit - The standard of measurement.
Statistic - A datum that can be represented numerically.
Term Accession Number - Identification number of a term selected from an ontology or a controlled vocabulary, and entered as a value for the Unit.
Term Source Ref - Name of the ontology or controlled vocabulary from which a term is selected and entered as a value for the Unit.
The Assay file will vary depending on the type of assay performed and protocol and technology type leveraged. Provided below are examples only of Assay files for some of the common types of assays identified across nanotechnology resources.
1.1.1 Example Assay file: Size by Dynamic Light Scattering (DLS)
Dynamic Light Scattering (DLS) is a type of spectroscopy that utilizes a laser beam to irradiate a sample containing particles in suspension, resulting in light scattering. Rapid fluctuations in scattering intensity, around a mean value at a certain angle, occur because of particle diffusion and are dependent upon on particle size. The calculated correlation function yields a diffusion coefficient, for a given temperature and viscosity, which can be used to calculate particle size. Exhibit 4.15 illustrates and example Assay file for Size by DLS.
Exhibit 4.15 Example Assay file for Size by DLS
1.1.2 Example Assay file: Zeta Potential
Zeta potential is a measurement of the magnitude of the electrical potential as generated by ion accumulation at the particulate surface. This surface consists of two layers, the Stern layer and the diffuse layer, which comprise an electrical double-layer. The measurement of the electrical potential provides important information regarding the dispersion mechanism, the degree of particle dispersion and the electrostatic potential of the sample. Exhibit 4.16 illustrates and example Assay file for Zeta Potential.
Exhibit 4.16 Example Assay file for Zeta Potential
1.1.3 Example Assay file: Assay file for Hemolysis
The hemolysis assay quantitates the amount of hemoglobin released from erythrocytes after treatment with a chemical or drug. Exhibit 4.17 illustrates and example Assay file for Hemolysis.
Exhibit 4.17 Example Assay file for Hemolysis
1.1.4 Example Assay file: Assay file for Cell Viability (MTT)
The MTT assay is a colorimetric assay that can assess the viability of cells by quantitation of the reduction of the yellow substrate MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) to a product that has a purple color. This assay can measure the cytotoxicity of a chemical or drug by determining the affect of treatment on cell viability. Exhibit 4.18 illustrates and example cell viability assay (MTT).
Exhibit 4.18 Example Assay file for Cell Viability (MTT)
1.1.5 Example Assay file: Assay file for Caspase 3 Apoptosis
Caspase 3 Apoptosis is an assay that assesses the activity of the enzyme caspase-3 to estimate the amount of apoptosis in a cell culture. Exhibit 4.19 illustrates and example caspase 3 apoptosis assay file.
Exhibit 4.19 Example Assay file for Caspase Apoptosis
1.1.6 Example Assay file: Assay file for Pharmacokinetics
Exhibit 4.20 illustrates and example assay file for pharmacokinetics.
Exhibit
