Haemolysis of sensitized erythrocytes (EA) can be carried out by incubating them successively with R2, followed by R1. When heated guinea-pig serum is present during the incubation of R2 with EA it inhibits haemolysis, though it has no effect when present during the incubation with R1. This inhibition is stronger with aged preparations of R2 which have much lower titres of C′4 than fresh preparations, though this increased inhibition does not occur because of the inactivation of C′4. Pre-incubation of EA with heated serum does not cause inhibition of their subsequent haemolysis by R2 and R1. Guinea-pig serum that has been decomplemented with an antigen-antibody precipitate and then heated is still inhibitory. Heated serum has no inhibitory action when haemolysis is carried out with whole guinea-pig serum in which all the components of complement are present simultaneously.
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