Immunochemical quantitaion of the third component of guinea pig complement in fluid phase and bound to cell surfaces.
A method is described for the determination of the absolute amount of guinea pig (GP) C-3 bound to cell surfaces. The test is based on the inhibition of anti-C-3 by C-3. The C-3 to be measured is added to a standard amount of anti-C-3 antibody and the residual activity is determined by its capacity to sensitize EC43-gp for lysis by GPC. By comparing the observed lysis with inhibition-of-lysis curves obtained with known C-3 antigen, the amount of test antigen can be determined. The method is simple, rapid, and reproducible and can detect nanogram amounts of C-3. The C-3 content of six normal guinea pig sera was found to be 1365 plus or minus 56 mug/ml (ranges 1120 to 1510 mug/ml) by the inhibition method described in this paper. Guinea pig hepatoma cells, line-1 and line-10, harvested from the peritoneal cavity of guinea pigs contained approximately 6.7 x 10-3 to 6.2 x 10-4 GPC-3 per cell whereas tumor cells sensitized with anti-Forssman or specific anti-tumor antibody contained from 1 to 9 x 10-5 C-3 per cell. Susceptibility to killing by antibody and GPC could not be correlated with the amount of GPC-3 bound to the surfaces of the tumor cells. Comparisons made between the amounts of C-3 bound to antibody-sensitized sheep erythrocytes and nucleated cells suggest there is a fundamental quantitative difference in the fixation of selected C components.
D000818:Animals; D000906:Antibodies; D002286:Carcinoma, Ehrlich Tumor / Q000276:immunology; D003165:Complement System Proteins / Q000032:analysis*; D004048:Diethylamines; D004912:Erythrocytes / Q000276:immunology; D005577:Forssman Antigen; D006168:Guinea Pigs / Q000276:immunology; D006461:Hemolysis; D007106:Immune Sera; D007111:Immunity, Cellular; D007700:Kinetics; D008113:Liver Neoplasms / Q000139:chemically induced / Q000276:immunology; D009603:Nitroso Compounds; D011817:Rabbits / Q000276:immunology; D012756:Sheep / Q000276:immunology
D000906:Antibodies; D004048:Diethylamines; D007106:Immune Sera; D009603:Nitroso Compounds; D003165:Complement System Proteins; D005577:Forssman Antigen