The establishment of a standard curve, which can be used to look up the CB1a concentration of a test sample from its measured adsorption.

<p>If an ELISA experiment is repeated with different solutions of known CB1a concentration; a calibration curve can be made, which shows the OD_450nm (A/A₀) values produced by different, given CB1a concentrations. Using this standard curve, an unknown concentration of CB1a in a test sample can then be found by finding its OD_450nm (A/A₀) value in an ELISA experiment. (<i>A</i>) The absorbance at 450 nm (OD_450nm) for different dilution rates (1∶2000; 1/10,000; 1/20,000) of secondary antibody (conjugated with horseradish peroxidase, HRP) and known concentrations of CB1a (ng/ml). (<i>B</i>) Log [CB1a] versus ELISA absorption (A/A₀); with 1/2000 dilution rate of secondary antibody used. The plotted line is y = −0.64x+0.5339. This is our calibration curve.