Functional identification of serum complement components following electrophoresis in polyacrylamide gels containing sodium dodecyl sulphate.
A method is described for detecting the active complement components C6 and C7 after polyacrylamide gel electrophoresis (PAGE) of whole serum in the presence of sodium dodecyl sulphate (SDS). The method involves the removal of SDS by washing with non-ionic detergent followed by the application of an erythrocyte/agarose gel to detect haemolytic activity. Two forms of human C6 with apparent molecular weights of approximately 121,000 daltons and 114,000 daltons were observed. Major activity resided in the 121,000 dalton species. The 2 forms of human C6 were not related to known genetic polymorphisms for this component. Analysis of sera from different animal species showed that not all possessed the 2 forms of C6 and that there were interspecies differences in C6 molecular weights. These are most marked in the case of human and murine C6; the major form of murine C6 had a molecular weight approximately 20,000 daltons less than the major human form. One form of human C7 with an apparent molecular weight of 104,000 daltons was seen. The molecular weights of C7 from the various animal sera tested did not differ significantly from this. Studies with reducing agents and metabolic inhibitors showed that both C6 and C7 required intact disulphide bonds and sulphydral groups for functional activity.
D000818:Animals; D003167:Complement Activation; D003183:Complement C6 / Q000276:immunology; D003184:Complement C7 / Q000276:immunology; D003165:Complement System Proteins / Q000276:immunology*; D004229:Dithiothreitol / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008623:Mercaptoethanol / Q000494:pharmacology; D012967:Sodium Dodecyl Sulfate; D013045:Species Specificity
D003183:Complement C6; D003184:Complement C7; D012967:Sodium Dodecyl Sulfate; D008623:Mercaptoethanol; D003165:Complement System Proteins; D004229:Dithiothreitol