Characterization of cell line β₃expression. (A) Flow cytometry analysis of CHO, 293, PMA-induced K562, HEL, Dami, HMEC-1, and WM793 cells. Fluorescence (FL) is shown on a logarithmic scale on the x-axis; cell number is shown on the y-axis. Dashed or dotted lines are the results using MoAb T10, which is specific for human _IIbβ₃ on CHO, PMA-induced K562, HEL, and Dami cells, or β₃-specific MoAb AP-3 for 293, HMEC-1, and WM793 cells; solid lines are the negative control using mouse Ig as the primary antibody. FACS analyses were performed at different times such that baseline fluorescence differed among cell lines. The key finding was the shift in fluorescence using MoAbs specific for β₃ or _IIbβ₃. (B) Northern blot analysis of 10 μg total RNA from selected cell lines probed with a full-length 2.6-kb β₃ cDNA and exposed to film at −80°C for 14 hours (upper panel). The β₃ mRNA signal in HEL cells is more easily seen on 24 hours of exposure. The filter was stripped and rehybridized with a GAPDH cDNA to assess loading equivalency (lower panel). Note that HEL and Dami cell β₃ RNA levels correlate well with protein expression, whereas PMA-stimulated K562 cells do not, suggesting that PMA-stimulation has complex effects on β₃ protein expression.