Matrix Interference in ELISA Experiments

Matrix interference (or matrix effects) is unwanted interactions between your protein of interest and other components in your sample. This can skew results and decrease the accuracy of your immunoassay, giving you lower than expected OD readings. These undesirable effects are more commonly observed in plasma and serum samples. 

Typically, matrix interference can be attributed to other components in the sample (such as phospholipids, carbohydrates, or proteins), high viscosity, salt concentrations, direct interactions between the protein of interest and other proteins in the sample, or the pH of the sample. 

If you think you might be experiencing matrix interference, you can run an experiment where a known amount of standard protein is added to a sample, known as “spiking”, and compare it to results obtained where the standard is diluted with dilution buffer. We suggest keeping the volume of the standard added to samples to a minimum to not disturb the matrix. If the readings do not match, you may be experiencing matrix interference. From here, you can calculate percent recovery with the following equation:

(Spiked Sample Concentration – Sample Concentration) / Spiked Standard Diluent Concentration

It would be ideal to have 100% recovery, but this is often not the case. A good recovery is anywhere from 80-120%. When matrix interference is present, your percent recovery calculated from the above equation will typically be less than 80%.

If you determined matrix interference is having an effect on your experiment, there are a few things you can do to try limit these effects: 

Dilution

Dilute your samples in sample dilution buffer so that the matrix effect can be diluted out. This may take some time to determine the appropriate dilution and will vary depending on samples and intensity of matrix interference. Ensure that you use the same buffer when diluting your samples for the standard curve and that your readings fall within the range of the assay. Don’t forget to take the dilution factor into account when calculating results.

Calibration

Dilute the standards and samples in the sample matrix. For example, if you are using serum samples, dilute both the standards and samples in normal serum. This may take some time to determine the appropriate dilution necessary to balance the effects.

If you think you may be experiencing matrix interference in your experiment, do not hesitate to contact us and we will discuss your results with our lab team to offer our feedback and suggestions!