Evaluating in-house anti-serum against B cells with flow cytometry

The applications of antibodies, be it monoclonal or polyclonal, in the diagnostic and research fields are well established. The disadvantage is the high cost of commercially available antibodies. In a diagnostic establishment like ours which also functions as a training ground for laboratory related personnel, it is beneficial to be able to produce in-house reagents. Therefore, we have undertaken this project to produce a rabbit polyclonal antibody against B lymphocytes. We found that the rabbit was a good choice because the titre of antibody produced was high and positive reactions were still detected at a dilution of 1:38400. The antibody showed significant positive reaction only with the lymphocyte subpopulation. A positive reaction was observed between the immunized rabbit serum and B lymphocytes but not T lymphocytes. This shows that the antibody was B lymphocyte specific. There was a positive correlation between the percentage of B lymphocytes labelled using the commercial anti-CD19 monoclonal antibody and the in-house polyclonal antibody (n = 13, r = 0.7, p = 0.02). However, the percentage of cells labelled by the in-house polyclonal anti-B was lower than that by the commercial monoclonal anti-CD19. The fluorescence intensity of the polyclonal antibody was lower than that of the monoclonal. In general, the performance of the in-house polyclonal antibody can be considered as satisfactory. The rabbit serum was stored at -20 degrees C and no significant loss of activity was detected for over a period of 19 months.