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ELISA Optimization Tips:

Antibody / Antigen Concentration

In order to get the best results from your ELISA assay, the dilution factors of the sample and the detection antibodies must be optimized. For strong, quantifiable signal, use a checkerboard titration to test for the optimal concentration of sample and detection antibodies.

How concentrated should the antibodies be?

In order to get the best results from your ELISA assay, the dilution factors of the sample and the detection antibodies must be optimized. If your sample or antibodies are too concentrated, you risk saturating the assay; if they are not concentrated enough, your signal will be weak and difficult to detect. For strong, quantifiable signal, use a checkerboard titration to test for the optimal concentration of sample and detection antibodies.

By running each well with a different ratio of sample to antibody, you can find not only the optimal concentration of each but the optimal ratio of concentrations as well. Using the information gleaned from the checkerboard assay, you can perform your ELISA experiment with the optimal concentrations for your application and get better results.

Uses of Checkerboard Titration

A checkerboard titration can be used to assess two variables at once: in this case, antibody concentration and sample concentration.

Checkerboard Titration

An example of a checkerboard titration. Each of the columns 1-12 contain the antibody dilution factors, and rows A-H contain sample dilution factors.

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