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The Effect of Benzylation or Sequential Amidination and Benzylation on the Ability of Rabbit G-Antibody to Fix Complement

Department of Immunochemistry, Walter Reed Army Institute of Research, Washington, D.C.

Abstract

Benzylation of rabbit G-antibody results in a partial loss of complement-fixing ability, as has been described by others. The greater part of this loss is due to the conjugation of only one or two tryptophan groups. The tryptophan groups which contribute most to the antibody's ability to fix complement are most susceptible to benzylation.

Double conjugation of rabbit G-antibody, by means of sequential amidination and benzylation, results in an antibody preparation which shows no significant loss in its ability to combine with and precipitate antigen, but which has suffered a 92% reduction in complement-fixing ability. This reduction is probably due to the specific inactivation of complement-fixing sites on the antibody molecule. Moreover, amidination and benzylation affect the same population of complement-fixing sites.

The doubly conjugated antibodies show a diminution in the rate at which they aggregate with antigen. However, double conjugation is more effective at maximizing changes in complement fixation and minimizing alterations in reaction with antigen than either amidination or benzylation alone.