An immunofluorescent technique for the detection of lymphocyte alloantigens

A triple-layer immunofluorescent technique for the detection of cell surface alloantigens has been developed. Chicken lymphoid cells were treated in sequence with: 1) chicken alloantibody, 2) rabbit anti-chicken Fc or anti-gamma chain serum and 3) FITC-labelled sheep anti-rabbit globulin. The second layer was diluted to that point where normal serum treated lymphocytes were no longer stained but without diminishing the degree of specific staining. This method enables the study of alloantigens expressed on B cells, avoiding the problem of staining of surface Ig-bearing lymphocytes.