Enzyme-linked immunosorbent assay (ELISA) is a method allowing the quantification of a desired marker in a biological sample. Achieving high quality quantification data by ELISA can be very advantageous when compared to more qualitative methods like IHC and Western blotting. For ELISA users, having a low coefficient of variability (CV or %CV) between sample replicates is crucial in demonstrating an assay was well-run and the resultant data are precise and accurate. Reliable assay results are assessed by standardized measures such as coefficient of variability.

        The coefficient of variability is a dimensionless numerical ratio used to describe the level of variability within a population independently of the absolute values of the observations. In statistical analysis of numerical data, if your absolute values are similar, sample populations can be assessed by using standard deviations; when absolute values vary, you must consider using a more standardized approach such as %CV, to assess the precision of a laboratory technique. CV is calculated by dividing the standard deviation (σ) of a set of measurements by the mean (µ) of the set which is then expressed as a percentage of variation to the mean (Figure 1).

         In ELISA data interpretation, %CV can highlight inconsistencies among sample replicates which is demonstrated in the data as variation among Optical Density (OD) readouts post-assay. These directly reflect the performance of the assay in the hands of the end-user. There are two types of %CVs that are used to express the precision of immunoassay results: intra-assay CV and inter-assay CV. Intra-assay CV is a measure of the variance between data points within an assay, meaning sample replicates ran within the same plate. Inter-assay CV is a measure of the variance between runs of sample replicates on different plates that can be used to assess plate-to-plate consistency. As a general guideline, to gauge the overall reliability of your immunoassay results, inter-assay %CV should be less than 15% while intra-assay %CV should be less than 10%.