PIGA^-RBCs are impaired in the regulation of autologous complement. (A) Increased in vitro C3 deposition on PIGA^- RBCs after activation of autologous complement. C3 deposition after complement activation induced by RBCs sensitized with various concentrations of antibody (classical pathway). C3 deposition is assessed measuring the mean fluorescence intensity (MFI) using a fluorescent monoclonal antibody against C3 and by flow cytometry. Data represent mean ± SD. RαM indicates rabbit antimouse RBC antibody. (B) PIGA^- RBCs are more susceptible to CVF-induced complement lysis in vivo. Wild type, FL, and GL mice were injected intraperitoneally with 5 mg/kg CVF. Free hemoglobin was measured by spectrophotometry at OD415 before and one hour after administration of CVF. Before injection (t = 0) the OD415 of plasma from wt, FL, and GL was not significantly different. One hour after plasma, OD415 measurements increased in all mice, but were significantly higher in FL and GL mice compared with the measurements in wt mice. Data represent mean ± SD.