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 consensus repeats (SCR), a Ser/Thr (ST)-rich region providing O-glycosylation sites, and the membrane-anchoring unit. A mAb, named D17, was raised against purified erythrocyte-DAF. This mAb recognized DAF on blood cells and most cell lines as determined by flow cytometry and immunoblotting. Its reactivity was similar to but weaker than that of two other 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-glycanase, suggesting that its epitope is located close to the O-glycosylation sites, probably within the ST-rich region or SCR4. D17 barely blocked the decay-accelerating activity of DAF. Using the three mAbs, tissue-associated and soluble forms of DAF were identified by SDS-PAGE/immunoblotting 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-associated and soluble forms of DAF, which were neither recognized by mAb against membrane cofactor protein (MCP, CD46) and C3b/C4b receptor (CR1, CD35) nor by non-immune IgG. In contrast to IA10 and 1C6, D17 did not recognize either spermatozoa-DAF or seminal plasma-DAF, or the deglycosylated or untreated forms of them. Immunohistochemical analysis showed that testis was stained with IA10 but not with D17.(ABSTRACT TRUNCATED AT 250 WORDS)
Hara
T
T
Department of Immunology, Center for Adult Diseases Osaka, Japan.
Matsumoto
M
M
Fukumori
Y
Y
Miyagawa
S
S
Hatanaka
M
M
Kinoshita
T
T
Seya
T
T
Akedo
H
H
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D015703:Antigens, CD / Q000276:immunology*; D018958:CD55 Antigens; D003429:Cross Reactions / Q000276:immunology; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000276:immunology; D004912:Erythrocytes / Q000276:immunology; D005434:Flow Cytometry; D006801:Humans; D007124:Immunoenzyme Techniques; D008214:Lymphocytes / Q000276:immunology; D008297:Male; D008562:Membrane Glycoproteins / Q000276:immunology*; D051379:Mice; D012661:Semen / Q000276:immunology*; D013094:Spermatozoa / Q000276:immunology; D013737:Testis / Q000276:immunology; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D018958:CD55 Antigens; D000939:Epitopes; D008562:Membrane Glycoproteins