Expression of CD15 (FAL) on myeloid cells and chromosomal localization of the gene

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Abstract
Different CD15 murine monoclonal antibodies were studied. These antibodies appeared to react specifically with the human myeloid-lineage-derived cell types in both peripheral blood and bone marrow. The antigens recognized by these antibodies were immunoprecipitated from lysates of125I-labelled neutrophilic PMNs of healthy donors and subsequently analysed by electrophoresis on SDS-polyacrylamide gel and autoradiography. All antibodies precipitated the same membrane polypeptides from the membrane-iodinated PMN lysates: 105 and 150-kDa as most prominent, together with 260-, 230-, 67- and 52-kDa polypeptides. Absorption studies were performed with synthesized carbohydrate molecules. Antibody B4.3 appears to be directed against 3-α-fucosyl-N-acetyl-lactosamine (FAL). Competition experiments with125I-labelled B4.3 demonstrated complete inhibition of binding by B4.3 and three other CD15 antibodies (VIM D5, UJ308, MI/N1), and partial inhibition by three additional antibodies (FMC10, FMC12, FMC13), indicating binding to the same antigenic structure. None of the antibodies reacted with monocytes using the immunofluorescence technique, but after neuraminidase digestion of these cells, positive reactions were obtained with all antibodies. Immunoprecipitation with lysates of both native and neuraminidase-digested monocytes showed no polypeptide bands. Monocytic differentiation of the myeloid cell line HL60 by 12-O-tetradecanoylphorbol-13-acetate (TPA) was accompanied by a decrease in reactivity with the antibodies, which could be reversed by neuraminidase digestion. This indicates that 3-α-fucosyl-N-acetyl-lactosamine is masked for the detection with antibodies upon monocytic differentiation by sialylation. Human x mouse myeloid cell hybrids were obtained after fusion of human myeloid cells and the HPRT-deficient murine myeloid cell line WEHI-TG. These hybrids were tested for reactivity with the anti-CD15 antibodies. The CD15 panel exhibited very similar reactivity patterns with the hybrid clones. Chromosomal analysis of hybrid cell metaphases revealed that the gene(s) involved in the expression of FAL must be located on human chromosome 11 in the region q12-qter.