A MONOCLONAL-ANTIBODY AGAINST HUMAN DECAY-ACCELERATING FACTOR (DAF, CD55), D17, WHICH LACKS REACTIVITY WITH SEMEN-DAF

Human decay-accelerating factor (DAF, CD55) is a phosphatidyl inositol -anchored glycoprotein consisting, from the N-terminus, of 4 short con sensus repeats (SCR), a Ser/Thr (ST)-rich region providing O-glycosyla tion sites, and the membrane-anchoring unit. A mAb, named D17, was rai sed against purified erythrocyte-DAF. This mAb recognized DAF on blood cells and most cell lines as determined by flow cytometry and immunob lotting. Its reactivity was similar to but weaker than that of two oth er well-characterized mAbs to DAF, IA10 (seeing an epitope within SCR1 ) and 1C6 (seeing an epitope within SCR3). The reactivity of D17 with erythrocyte DAF became increased by treatment with sialidase/O-glycana se, suggesting that its epitope is located close to the O-glycosylatio n sites, probably within the ST-rich region or SCR4. D17 barely blocke d the decay-accelerating activity of DAF. Using the three mAbs, tissue -associated and soluble forms of DAF were identified by SDS-PAGE/immun oblotting and immunohistochemical staining. IA10 and 1C6 recognized a 50 kDa protein in spermatozoa lysate and two proteins of Mr 70 and 55 kDa, respectively, in seminal fluid. These represented membrane-associ ated and soluble forms of DAF, which were neither recognized by mAb ag ainst membrane cofactor protein (MCP, CD46) and C3b/C4b receptor (CR1, CD35) nor by non-immune IgG. In contrast to 1A10 and 1C6, D17 did not recognize either spermatozoa-DAF or seminal plasma-DAF, or the deglyc osylated or untreated forms of them. Immunohistochemical analysis show ed that testis was stained with IA10 but not with D17. These findings indicate that DAF in seminal plasma and on spermatozoa are unusual in that the D17 epitope is lacking or inaccessible.