Comparative analysis of cell surface markers on murine NK cells and CTL target-effector conjugates

Abstract

The rosetting of sheep erythrocytes (SRBC) coated with non-haemagglutinating monoclonal antibodies rather than conventional haemagglutinating antisera revealed readily detectable FcR on most splenic natural killer (NK) cells since 76% of splenic lymphocytes forming conjugates with YAC also rosetted with SRBC coated with high concentrations of monoclonal anti-SRBC antibody of the IgG2b subclass and since Ficoll depletion or enrichment of splenic lymphocytes rosetting with IgG2b-coated SRBC resulted in a corresponding 4-fold decrease or increase in conjugate-forming cells and a 10-fold decrease or increase in NK cytolytic activity. NK cells bound much less readily to monoclonal IgG2a and not at all to monoclonal IgG1 or IgM, but the degree of binding was directly proportional to the amount of antibody on the erythrocytes and was not isotope-restricted. In addition, immunofluorescent studies revealed that YAC-1-conjugated lymphocytes were Lyt-1-, Lyt-2-, partially Thy-1+ (60%), asialo(GM1+ (80%). Qa-4+ (77%), Qa-5+ (79%), and Ly-5+ (94%). In comparison, a proportion (39%) of alloimmune peritoneal exudate cells which conjugated with P815-2 also stained by immunofluorescence with anti-asialo GM1 antisera. Most (greater than 90%) P815-conjugated cells were Thy-1+, Lyt-2%, and a subpopulation of Lyt-1+2+ conjugates was observed (25%). Qa-5 and Ly-5 were also expressed on most (two-thirds) cytolytic T lymphocytes (CTL) conjugates, whereas Qa-4 and FcR for IgG2b were not detected. The best phenotype distinctions between NK cells and CTL were therefore based on the presence or absence of Lyt-2, Qa-4, and FcR for IgG2b on most effector cells. Anti-asialo-GM1 or monoclonal anti-Qa-4 and complement treatment greatly diminished both the frequency of NK conjugates and the percentage of conjugates with detectable IgG2b FcR or asialo-GM1. These results confirm that NK cells co-express asialo-GM1 and Fc receptors, at the single-cell level, and provide a simple method for greatly enriching NK populations at least 10-fold.