A monoclonal antibody that blocks the complement regulatory activity of guinea pig erythrocytes and characterization of the antigen involved as guinea pig decay-accelerating factor.
MCA44 is a mAb with the capacity to sensitize neuraminidase-treated guinea pig E for hemolysis by homologous guinea pig C, and the Fab fragments of this mAb could also sensitize guinea pig E interfering with the function of a membrane inhibitor of C on guinea pig E. Using an immunosorbent column to which MCA44 was coupled, the antigenic molecule termed 44Ag was purified from the glycoprotein fraction extracted from E membranes. C intermediate sheep E treated with guinea pig C1 and C4 after sensitization with Ab (EAC14b cells) lost the ability to generate C3 convertase with C2 after incubation with 44Ag. Treatment of guinea pig E and PBL with phosphatidyl-inositol specific phospholipase C (PIPLC) partially removed 44Ag, as determined by flow cytometric analysis after immunofluorescence staining with MCA44. However, 125I-labeled 44Ag adsorbed to human E was efficiently removed by PIPLC treatment with a slight reduction in M(r). The 44Ag purified on an immunosorbent column showed three bands on SDS-PAGE. However, partial N-terminal amino acid sequences of the 55-kDa, 70-kDa, and 88-kDa bands under nonreducing conditions were identical and the sequence was 55% homologous to the N-terminal sequence of human decay accelerating factor (CD55). Intracutaneous administration of MCA44 or its F(ab')2 fragment resulted in increased capillary permeability, even after 3 days, as determined by the appearance of Evans blue spots after i.v. administration of the dye. Because control Abs including anti-class I-MHC did not cause such increased capillary permeability, the increase in permeability caused by MCA44 was likely induced by blocking the function of 44Ag in vivo, indicating a crucial role for these molecules in preventing over-activation of C at the site.
Okada
N
N
Department of Molecular Biology, Nagoya City University School of Medicine, Japan.
Tanaka
H
H
Takizawa
H
H
Okada
H
H
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D015703:Antigens, CD / Q000737:chemistry* / Q000276:immunology*; D015153:Blotting, Western; D018958:CD55 Antigens; D003169:Complement Inactivator Proteins / Q000737:chemistry / Q000276:immunology*; D004912:Erythrocytes / Q000737:chemistry / Q000276:immunology; D005434:Flow Cytometry; D006168:Guinea Pigs; D007249:Inflammation / Q000276:immunology / Q000473:pathology; D008081:Liposomes / Q000032:analysis; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000276:immunology*; D008969:Molecular Sequence Data; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D010727:Phosphoric Diester Hydrolases / Q000502:physiology
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D018958:CD55 Antigens; D003169:Complement Inactivator Proteins; D008081:Liposomes; D008562:Membrane Glycoproteins; D010727:Phosphoric Diester Hydrolases; D043265:Phosphatidylinositol Diacylglycerol-Lyase