For these samples, the concentration obtained from the standard curve when analyzing the results must be multiplied by the dilution factor. To obtain an accurate result, these samples should be diluted or concentrated before proceeding with the ELISA staining. Samples that have an absorbance value falling out of the range of the standard curve The concentration of the positive control sample should be within the linear section of the standard curve in order to obtain valid and accurate results. We recommend using a sample of known concentration as a positive control. Each point on the graph represents the mean of the three parallel titrations. We recommend including a standard on each ELISA plate to provide a standard curve for each plate used.Ī representative standard curve is shown in the figure below, from human HIF1 alpha SimpleStep ELISATM kit (ab171577). Draw a best fit curve through the points in the graph (we suggest that a suitable computer program be used for this). Duplicates should be within 20% of the mean.Ĭreate a standard curve for the target protein by plotting the mean absorbance (y axis) against the protein concentration (x axis). Many computer programs are now available to help process ELISA results in this way.Ĭalculate the average absorbance values for each set of duplicate standards and duplicate samples. This will provide enough data for statistical validation of the results. Always run ELISA samples in duplicate or triplicate.

Elisa curve on graphpad prism