Journal of Cell Science, Vol 78, Issue 1 117-131, Copyright © 1985 by Company of Biologists

JOURNAL ARTICLES

Enrichment of CFU-S proliferation inhibitor-producing cells based on their identification by the monoclonal antibody F4/80

PJ Simmons and BI Lord

Cells capable of synthesizing a factor that can specifically inhibit the proliferation of haemopoietic spleen colony-forming units, CFU-S, carry many of the characteristics of macrophages. The monoclonal antibody, F4/80, which is specific for murine macrophages, has therefore been used to isolate macrophages from bone marrow. These macrophages were then assayed for their ability to produce the inhibitor. Low-density bone marrow cells were first separated by a density-cut procedure and then labelled with F4/80. Fluorescence-activated cell sorting was then used to select F4/80 positive and negative cell fractions. It was found that the F4/80 positive fraction contains inhibitor cells, enriched by at least 200-fold compared to unfractionated marrow. Subsequent culture of this fraction over a period of several weeks produced a further 15- to 20-fold increase in inhibitor-producing capacity. The cultured cells producing inhibitor were virtually 100% F4/80 positive, phagocytic and exhibited histochemical properties characteristic of macrophages. The macrophage-like character of the producer cells was thus confirmed and the removal of the majority of unwanted cells means that many of the impurities in the normal crude extracts have been excluded.