Single-step purification and evaluation of recombinant BP26 protein for serological diagnosis of Brucella ovis infection in rams.
To investigate the value of the BP26 protein in the serological diagnosis of ovine brucellosis caused by Brucella ovis, recombinant BP26 protein was produced in Echerichia coli and purified for use in an indirect enzyme-linked immunosorbent assay (I-ELISA). The majority of the recombinant protein was recovered from the supernatant of sonicated recombinant E. coli cells in a soluble form. This facilitated the purification of the recombinant BP26 protein which was achieved by using ion-exchange chromatography. After one step of purification, the purity of the recombinant BP26 protein was analyzed by using SDS-PAGE, Coomassie blue staining, and Western blot with a monoclonal antibody (MAb) directed against the BP26 protein. The degree of purity appeared satisfactory so that it could be directly used in I-ELISA. Although the recombinant BP26-ELISA appeared less useful than I-ELISA using the B. ovis hot saline (HS) extract as antigen, the high number of sera from B. ovis infected rams found positive (90%) in the recombinant BP26-I-ELISA indicated that the BP26 protein may be an additional suitable antigen for serological diagnosis of B. ovis infection in rams.
Zygmunt
Michel S
MS
Laboratoire de Pathologie Infectieuse et Immunologie, Institut National de la Recherche Agronomique, F-37380 Nouzilly, France. zygmunt@tours.inra.fr
Baucheron
Sylvie
S
Vizcaino
Nieves
N
Bowden
Raul A
RA
Cloeckaert
Axel
A
D000818:Animals; D000907:Antibodies, Bacterial / Q000097:blood; D015153:Blotting, Western / Q000662:veterinary; D002002:Brucella / Q000302:isolation & purification*; D002006:Brucellosis / Q000175:diagnosis / Q000382:microbiology / Q000662:veterinary*; D002852:Chromatography, Ion Exchange / Q000662:veterinary; D004591:Electrophoresis, Polyacrylamide Gel / Q000662:veterinary; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000662:veterinary*; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008297:Male; D008565:Membrane Proteins / Q000302:isolation & purification*; D011994:Recombinant Proteins / Q000302:isolation & purification; D012756:Sheep; D012757:Sheep Diseases / Q000175:diagnosis / Q000382:microbiology*
D000907:Antibodies, Bacterial; C106040:BP26 protein, Brucella; D008565:Membrane Proteins; D011994:Recombinant Proteins