Representatives of two species, the chicken and the goat, were immunized by injection with Freund's adjuvant, of specific precipitates prepared from the antisera of rabbits with various allotypic specificities. The precipitation reactions of the resulting antisera with normal rabbit sera were studied in liquid and in gellified media. In these reactions the isotypic specificity and often one or the other of the allotypic specificities of rabbit γ-globulin were involved.
It was always easier to obtain antibodies against the specificities controlled by the alleles of locus b (Ab4, Ab5, Ab6) than of locus a (Aa1, Aa2, Aa3). The precipitation reaction of antibodies against a specificity of the a series could always be inhibited by an excess (nearly always moderate) of a serum lacking this specificity. A similar inhibition of the antibodies against the specificities of the b series was often impossible, especially if the antiserum had been collected after a second injection of immunizing material.
The reactions of 58 Ab4+ sera with a goat anti-Ab4 serum and of 28 Ab5+ sera with a chicken anti-AbS serum were studied in gel tubes (simple diffusion). The constant observation of two precipitation zones due respectively to two types of molecules, Ab4+ and Ab4- (or Ab5+ and Ab5-), indicated the existence of an appreciable number of Ab4- or Ab5- molecules, even in supposed homozygotes. In the latter rabbits Ab4- or Ab5- molecules are b- molecules; i.e., molecules lacking the specificities of the b series. Measurements of the distances between the two zones in the reaction of 72 sera revealed significant differences between sera of individuals of the same phenotype, as well as systematic differences not only between sera of homozygotes and of heterozygotes, but also between heterozygotes of different genotypes. These measurements were used to evaluate the ratio of concentrations of the two types of molecules detected by each of the two antisera.
The occurrence of b- molecules in every one of the fairly large number of sera studied indicates a new aspect of the heterogeneity of γ-globulin. Although detected by means of allotypy, this heterogeneity is not dependent upon factors which vary with individuals.
These results are discussed in connection with what is known of the structure of γ-globulin.