EFFICIENT IMMUNOSELECTION OF CYTOLYTIC EFFECTORS WITH A MAGNETIC CELLSORTER

This paper describes a rapid and efficient method for the sorting of i n vitro activated cytolytic effectors cells. For cytotoxic assays, a l arge number of cells with conserved function must be rapidly obtained. Immunomagnetic sorting was chosen because it is faster than flow cyto metry sorting. The MACS system requires the use of paramagnetic beads of small diameter (100-150 nm), reputed to interfere minimally with ce ll function. In order to generate the cytolytic effectors, peripheral blood lymphocytes were cultivated in the presence of interleukin-2 (50 U/ml) and anti-CD3 monoclonal antibody (BMA030, 100 ng/ml) for 4 days . Cell separation was based on the membrane expression of the CD3 comp lex. The purity obtained for positive (CD3+) cell sorting with the MAC S was higher than 95 %. The purity of negative (CD3-) cell fraction wa s more variable, but further purification by flow cytometry rapidly yi elded purity higher than 95 %. Cytotoxic assays were performed against four target cell lines (K562, Daudi, HL60 and U937) and proliferation assays showed that both negatively and positively selected population s had conserved their function acquired during culture in the presence of anti-CD3 mAb and IL2.