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United States Department of Agriculture

Agricultural Research Service

Title: Isolation of Antigenic Outer Membrane Proteins from Brucella Abortus Rb51 by Lectin Affinity Chromatography

Authors
item Brees, Dominique - IA STATE UNIV
item Stevens, Mark
item OLSEN, STEVEN
item Cheville, Norman - IA STATE UNIT

Submitted to: Research Workers in Animal Diseases Conference Proceedings
Publication Type: Abstract Only
Publication Acceptance Date: November 14, 1995
Publication Date: N/A

Technical Abstract: Brucella abortus strain RB51 is being tested as vaccine for brucellosis in cattle because it does not induce production of lipopolysaccharide antibodies which are detected by serodiagnostic tests for brucellosis in vaccinated herds. However, the antibody response to strain RB51 has not been fully characterized and there is no serologic test which detects antibodies which are specific for RB51 in RB51-vaccinated cattle. This paper describes the isolation of diagnostic immunoreactive outer membrane proteins (OMP) from strain RB51 based on their affinity for binding to lectins. Strain RB51 OMP were separated by SDS-PAGE and reacted with seven different lectins. Of these seven, Con A had the highest reactivity with the OMP and this lectin was subsequently used to isolate immunoreactive OMP. Strain RB51 OMP were separated by isoelectric focusing and those with an isoelectric point of 4.11 to 4.42 were further purified by Con A affinity chromatography. The eluted Con A-binding proteins were analyzed by western blotting using antiserum from 8 strain RB51-vaccinated cattle. Nine proteins were identified which reacted with all tested antiserums. These results indicate that Con A affinity chromatography permitted the isolation of nine antigenic OMP from RB51. One or more of these proteins may be useful candidate antigens in developing a specific test for detecting antibody to RB51 in RB51-vaccinated cattle. Of the nine reacting proteins, one protein (18 kDa) is currently being further characterized by amino acid sequencing.

Last Modified: 7/25/2014
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