The preparation of antimacrophage serum (AMS) and its application to murine fibrous histiocytoma are reported. AMS, prepared by inoculating peritoneal macrophages from mice into rabbits, was purified by repeated absorption with kidney homogenate, red blood cells, thymus cells, non-adherent spleen cells and finally, L-cells. The molecular weight was determined as 170,000 daltons using SDS-polyacrylamide gel electrophoresis. AMS diluted to 1:32 reacted with 61.4% of peritoneal macrophages, but not red blood cells, lymphocytes, granulocytes and fibroblasts, as judged by membrane immunofluorescence. 57.1% of monoclonal B-10 macrophages and 75.6% of 28-12 macrophages transformed with SV40 reacted with AMS. B-10 and 28-12 cell tumors simulating human malignant fibrous histiocytoma gave a distinctive diffuse pattern of immunofluorescence with AMS which affected both round- and spindle-shaped cells, although positive fluorescence of spindle-shaped cells was somewhat weaker than that of round-shaped cells. However, tumor tissues from a murine fibrosarcoma showed no reaction with AMS. Based on these findings, AMS may be useful in determining the histogenesis of some soft tissue tumors of doubtful origin.