Improvement of the reproducibility of the ELISA test for anti- Trypanosoma congolense antibody-detection in cattle

Abstract

Simplicity and the potential automatization make the ELISA test a universal tool for the detection of antibodies, and, more recently, of antigens. But the reproducibility of results is not very good, due to many varying factors. We tried to improve the reproducibility of the ELISA test for the detection of anti-Trypanosoma congolense antibodies in cattle. For that, buffers are always used at room temperature to avoid temperature gradients in the plates. All volumes are increased to 200 µl per well. Non-activated rabbit serum is used to block non-specific sites and to reduce background signal. Better results were obtained with a homologous antigen (T. congolense) than with a heterologous one (T. evansi). Titration of the antigen concentration to be used must be checked for each new batch. A distribution is proposed which would permit a testing of each serum four times per plate and thus to calculate the mean and standard deviation for each serum. Finally, we propose that the reading should no longer be a function of time after contact between enzyme and substrat, but a function of the progress of the reaction measured in a target serum. Results show a very good reproducibility and in addition we propose a computerized monitoring system.