The first complement component: evidence for an equilibrium between C1s free in serum and C1s bound in the C1 complex

Abstract

An equilibrium between free C1s and C1s bound in macromolecular C1 exists in human serum. This equilibrium can be utilized to incorporate radioiodinated C1s into serum C1. Human sera were incubated for 40 hr at 4 degrees C with 125I-C1s to allow the exchange between free and bound C1s to reach equilibrium. The C1 complex labeled in this manner was separated from the majority of serum proteins by centrifugation in linear 10 to 30% sucrose density gradients. The resulting fractions containing 125I-C1 can be used directly and conveniently in C1 activation assays that detect the cleavage of proenzyme C1s. Electrophoretic analysis on polyacrylamide gels showed the presence of only proenzyme 125I-C1s in serum C1, whether or not the applied labeled material contained 125I-C1s or other labeled proteins. The inability of C1 to incorporate C1s was shown to be the result of decreased stability of C1 upon activation.