OPTIMIZING THE DETECTION OF CELL-SURFACE ANTIGENS ON ELICITED OR ACTIVATED MOUSE PERITONEAL-MACROPHAGES

Blocking conditions that are optimal for the detection of surface anti gens on resident peritoneal macrophages (PM phi) by flow cytometry are not ideal for elicited or activated PM phi. A blocking step of 10% go at serum can be used routinely to detect the F4/80 and Mac-1 antigens on resident PM phi. In contrast, high concentrations (33-50% each) of combined goat and mouse sera were required to reduce nonspecific bindi ng and to improve the detection of the F4/80 antigen on PM phi elicite d by thioglycollate broth (TG) or activated by maleic anhydride diviny l ether copolymer (MVE-2). However, even low concentrations of goat se rum masked the expression of the Mac-2 antigen on TG and MVE-2 PM phi. Thus, within a given elicited or activated PM phi population, differe nt blocking conditions may be necessary to detect different surface an tigens optimally. In addition to blocking, the use of isotypic control s that match the monoclonal antibody isotypes was found to be necessar y for the optimal detection of antigen expression on TG and MVE-2 PM p hi. (C) 1994 Wiley-Liss, Inc.