[Differentiation of leukocytes in thrombocyte concentrates by APAAP staining]

The preparation of platelet concentrates (PC) from pooled buffy coats has raised concern as to whether activation of lymphocytes might take place during the period of storage. Therefore it appears to be essential to investigate the degree of leukocyte contamination and identify the subtypes of these cells. Especially lymphocytes are of relevance as they may induce either a mixed lymphocyte reaction (MLR), the production of cytokines or a graft-versus-host reaction. As the number of leukocytes is very low we adopted the alkaline phosphatase-antialkaline phosphatase (APAAP) technique to determine the composition of leukocytes. We used mouse monoclonal antibodies to specifically stain T lymphocytes, B lymphocytes, monocytes, HLA-DR-positive cells and NK cells. All subtypes could easily be identified and their relative amounts were determined. In samples from 15 PCs the percentages of T lymphocytes, B lymphocytes, monocytes, NK cells and HLA-DR-positive cells was 40.3, 6.4, 18.9, 7.2 and 17.1% of total leukocytes, respectively. The standard deviation ranged between 2 and 5%. We highly recommend this technique, which is a very sensitive method to determine leukocyte contamination in PC.