Identification of mouse T and B lymphocytes from cytocentrifuged cell smears

A method enabling direct enumeration of mouse T and B cells from cytocentrifuged cell smears is described. The acid alpha-naphthyl acetate esterase (ANAE) staining was used as marker for T cells. The distribution of the ANAE marker-carrying lymphocytes closely followed the percentual distribution of T cells in spleen and lymph node. Electrophoresis analysis demonstrated that while more than 95% of resting spleen and lymph node T cells carried the ANAE marker, only less than 5% of the B cells were ANAE positive. The B cells (surface-bound Ig-carrying small lymphocytes) were identified by anti-Ig serum followed by rosetting of the B cells with Staphylococcus aureus strain Cowan 1 (StaCw). The ANAE marker-carrying cells did not overlap with the StaCw rosette-forming lymphocytes. Thus we conclude that the combination of StaCw rosetting and ANAE staining enables accurate identification of resting T and B cells from a single microscope slide. Ninety to 100% of Con A-activated blasts expressed the ANAE marker but only 60-85% of PHA and MLC-activated blasts were positive. Twenty to 33% of the blast cells stimulated by LPS expressed the ANAE marker. Thus the ANAE marker is not a reliable criterion for T cells in actibated state.