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.

Abstract

Abstract 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.