Characterization of mouse DAF on transfectant cells using monoclonal antibodies which recognize different epitopes.
Several membrane proteins prevent host cells from homologous complement attack. In humans, one such protein, decay-accelerating factor (DAF), exists as two isoforms, a GPI anchored form and a secreted form, which are generated by alternative splicing. DAF in mouse is also expressed as two isoforms, a GPI anchored form (GPI-DAF) and a transmembrane form (TM-DAF), which are produced from two separate genes. In this study, we transfected cDNA of mouse GPI-DAF or TM-DAF into Chinese hamster ovary (CHO) cells. Both isoforms of DAF on CHO cells were shown to regulate mouse complement C3 deposition mediated by the classical and alternative pathways and the inhibitory activity of both isoforms was species restricted. The two mouse DAF isoforms were effective against rat complement but not against human and guinea pig complement. Furthermore, we produced hamster mAbs to mouse DAF using GPI-DAF transfectant cells and established seven unique mAbs (RIKO-1-7). Western blotting analysis using RIKO-3, which reacts with both GPI-DAF and TM-DAF, and RIKO-4, which is an anti-GPI-DAF specific mAb, indicated that GPI-DAF was expressed on erythrocytes, spleen and testis, and that TM-DAF was expressed only in testis.
Ohta
R
R
Department of Molecular Biology, Nagoya City University School of Medicine, Aichi, Japan.
Imai
M
M
Fukuoka
Y
Y
Miwa
T
T
Okada
N
N
Okada
H
H
D000818:Animals; D000911:Antibodies, Monoclonal; D015153:Blotting, Western; D018958:CD55 Antigens / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D016466:CHO Cells; D003176:Complement C3 / Q000378:metabolism*; D006224:Cricetinae; D000939:Epitopes; D004912:Erythrocytes / Q000166:cytology / Q000378:metabolism; D017261:Glycosylphosphatidylinositols / Q000737:chemistry; D006168:Guinea Pigs; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D020033:Protein Isoforms / Q000378:metabolism; D051381:Rats; D013045:Species Specificity; D013154:Spleen / Q000166:cytology / Q000378:metabolism; D014162:Transfection; D010738:Type C Phospholipases / Q000378:metabolism
D000911:Antibodies, Monoclonal; D018958:CD55 Antigens; D003176:Complement C3; D000939:Epitopes; D017261:Glycosylphosphatidylinositols; D020033:Protein Isoforms; D010738:Type C Phospholipases; D043265:Phosphatidylinositol Diacylglycerol-Lyase