An immunoperoxidase method for the identification of B lymphocytes by light microscopy

Peroxidase-conjugated anti-human immunoglobulin was used to determine the percentage of B lymphocytes in the mononuclear cell fraction of peripheral blood of 100 healthy persons (blood donors). Peroxidase activity was revealed by incubation with the usual mixture of 3'3 diaminobenzidine and hydrogen peroxide. Cells which were peroxidase-positive after incubation with benzidine solution alone were considered to be monocytes. The number of T lymphocytes was estimated by the formation of E rosettes. 19.6% mononuclear cells could be shown to be B cells, 4.1% were monocytes and 57.0% T cells. The use of peroxidase-conjugated anti-human immunoglobulin gives rise to the same percentage of B cells as the use of fluorescein-conjugated anti-human immunoglobulin. The advantage of the method described here is that B lymphocytes can be counted by conventional light microscopy.