Veterinary Microbiology, 9 (1984) 549--560 549 Elsevier Science Publishers B.V., Amsterdam -- Printed in The Netherlands CHARACTERIZATION OF SALT-EXTRACTABLE PROTEIN ANTIGENS FROM BR UCELLA ABOR TUS BY CROSSED IMMUNOELECTRO- PHORESIS AND ISOELECTRICFOCUSING L.B. TABATABAI and B.L. DEYOE National Animal Disease Center, Agricultural Research Service, Science and Education, U.S. Department of Agriculture, P.O. Box 70, Ames, IA 50010 (U.S.A.) (Accepted 7 May 1984) ABSTRACT Tabatabai, L.B. and Deyoe, B.L., 1984. Characterization of salt-extractable protein antigens from Brueella abortus by crossed immunoelectrophoresis and isoelectro- focusing. Vet. Microbiol., 9: 549--560. Salt-extractable protein antigens (CSP) from Brucella abortus strains 19 and 2308 (vaccine and virulent strains, respectively) were analysed by crossed immunoelectro- phoresis (CIE) using rabbit antisera to protein antigens and by isoelectricfocusing (IEF) in polyacrylamide gels. The reference immunoelectrophoretic profiles developed for proteins from strain 19 and 2308 of B. abortus contained 20 and 25 immunoprecipitates, respectively. Serum from cows experimentally infected or hyperimmunized with live organisms produced up to 5 immunoprecipitates in CIE with the protein antigens. Ab- sorption of rabbit sera with homologous B. abortus cells reduced, but did not eliminate all of the immunoprecipitates from rabbit sera, suggesting that the majority, but not all of the protein components, are exposed on the surface of the cell. In contrast, antibody to protein antigens in agglutin-free absorbed serum from infected cattle could still be demonstrated by CIE, even though CIE with protein extracts from whole cells radio- iodinated with the cell surface labeling reagent, diazoiodusulfanilic acid, indicated that these antigens may be at or near the surface of the cell. From CIE in heterologous sys- tems we concluded that all proteins present in strain 19 preparations were partially or completely identical to those in strain 2308. The IEF studies paralleled the CIE studies and revealed that the protein profile from strain 2308 was more complex than the profile from strain 19. Major differences between the 2 strains were found in the pH region from 3.9 to 5.0, where strain 2308 exhibited 4 additional protein bands. INTRODUCTION Surface components of Brucella organisms have been shown to contain lipopolysaccharide (LPS) (Olitzki, 1959; Schurig et al., 1978), and numer- ous non-LPS antigenically distinct components detectable with sera from infected and vaccinated cattle (Corbel, 1973; Hinsdill and Berman, 1967; Schurig et al., 1978; Stemshorn and Nielsen, 1977; Tabatabai et al., 1979). In contrast, Group 2 and Group 3 outer-membrane proteins of B. abortus 0378-1135/84/$03.00 © 1984 Elsevier Science Publishers B.V.