Gen5 version required: 1.0 or higher
Basis for the Assay:
In a typical competitive binding assay the primary measure of the dose-response curve is the IC50 or the concentration that provides 50% inhibition. In this example, it is first necessary to calculate the theoretical % toxicity of the samples by dividing the mean blanked sample ODs by the mean blanked control (CTRL) ODs for each sample, then multiplying by 100. The resulting % toxicity is provided as the “Data In” to the 4-Parameter (4P) curve fit, with the requested interpolation set to 50. The concentrations of the samples are calculated from the curve and the interpolated 50% value is provided.
The protocol calls for an endpoint absorbance measurement at 620 nm. Background absorbance of the microplate is removed by subtraction of blank wells. Two dose-response curves are plotted based on 6 sample-dilutions, which along with two controls are pipetted to the plate in triplicate.
The experiment file contains the absorbance measurements used to determine theoretical toxicity and to plot the dose-response curves.
Two samples in dilutions of 10-60, each with a specific control and blank in triplicate.
The report contains a matrix with: 1. Well ID 2. Dilution value 3. Raw absorbance value 4. Calculated theoretical “Toxicity %.” The report also contains the two curves and their interpolations tables.