Summary: Antigens prepared from 22 genetically different smooth strains of Brucella were compared and gave similar agglutination titers with serums from established brucellosis and cross-reacting serums from cases of tularemia and from persons immunized against cholera. The temperature of incubation of the titrations and the method used for killing the antigen suspensions influenced the titers, especially in titrations of cross-reacting serums. Titers of cross-reacting serums with phenol-killed antigen suspension were much lower when titrations were incubated at 56 C than when they were incubated at 45 C or below; titers of brucellosis serums were essentially independent of incubation temperatures from 24 C to 60 C. The lower titers of cross-reacting serums at 56 C appeared to be due to a reversible inhibition of the non-specific agglutination. Titers with formalin-killed antigen suspension were influenced much less by incubation temperature; these antigens, however, favored the appearance of prozones, particularly with serums from recovered rather than from active brucellosis. The authors are indebted to Dr. Werner Braun for helpful discussion and comment during the course of the work, and for providing the Brucella strains.