Assessment of complement activation in clinical samples. Comparison of immunochemical and functional measurements of complement components with quantitation of activation fragments.
We have compared functional and immunochemical measurements of complement components with assays measuring the generation of activation fragments, for the assessment of classical pathway activation in vitro and in vivo. The generation of the C3a, C3b/C3bi cleavage fragments of C3, and of the C4d cleavage fragment of C4 measured by ELISA and RIA, was correlated with the decrease in total complement hemolytic activity (CH50) and in functional activity of C3 and C4 in normal human serum in which the classical pathway had been activated with aggregated IgG. The decrease in CH50 in in vitro activated serum was also correlated with the generation of C5a and soluble SC5b-9 complexes. In contrast, no or little increase in the concentration of C3a, C3b/C3bi and C4d was observed in plasma samples from patients with low CH50 and with low levels of immunochemical C3 and C4, indicating that fragment quantitation assays provide no information on the presence and extent of complement activation in vivo. Analysis of samples from patients expressing the four C4 genes and patients having one or two C4 null alleles indicated that a ratio of hemolytic C4 to C2 > or = 1 was indicative of complement activation without C4 deficiency, whereas a ratio below 1 was indicative of C4 deficiency with or without classical pathway consumption. Classical pathway activation and C4 deficiency in clinical samples are best predicted by the concomitant assessment of immunochemical levels of C3 and C4 and hemolytic levels of C4, C2 and C3.
Maillet
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Service d'Immunologie, Hôpital Broussais, Paris, France.
Frémeaux-Bacchi
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V
Uhring-Lambert
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B
Kazatchkine
M D
MD
D003167:Complement Activation; D003175:Complement C2 / Q000032:analysis*; D003176:Complement C3 / Q000032:analysis; D003181:Complement C4 / Q000032:analysis* / Q000172:deficiency / Q000235:genetics; D015936:Complement C5a / Q000032:analysis; D015941:Complement Hemolytic Activity Assay; D006461:Hemolysis; D006801:Humans; D007118:Immunoassay
D003175:Complement C2; D003176:Complement C3; D003181:Complement C4; D015936:Complement C5a