Studies of Culture-Derived Soluble Babesia Bovis Antigens: Purification, Characterization and Application in Serodiagnosis
Montenegro Heredia, Sonia Armonia
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https://hdl.handle.net/2142/71295
Description
Title
Studies of Culture-Derived Soluble Babesia Bovis Antigens: Purification, Characterization and Application in Serodiagnosis
Author(s)
Montenegro Heredia, Sonia Armonia
Issue Date
1982
Department of Study
Veterinary Medical Science
Discipline
Veterinary Medical Science
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Biology, Veterinary Science
Abstract
Analytical purification of soluble antigens from crude supernatant fluids of Babesia bovis cultures was conducted by various immunoadsorption procedures using horse anti-B. bovis IgG. A Protein A-Sepharose CL-4B adsorbent proved the most efficient as 58 mg of antigen could be purified from 34 ml of culture fluid. Isolated antigens were identified by crossed immunoelectrophoresis.
Subsequent preparative purification of three soluble babesial antigens was performed with ammonium sulfate precipitation and anion exchange chromatography. The antigens were precipitated with 60-70% saturated ammonium sulfate. Also, a 0.1M Tris-HCl buffer pH 6.5 selectively eluted the Babesia antigens from DEAE-cellulose with minimal serum contamination.
Monospecific rabbit antibodies produced against individual B. bovis antigens were used in an immunofluorescence test to identify the location of the respective antigens within the infected erythrocyte. Two antigens were found on or near the erythrocyte membrane, while a third antigen was associated directly with the parasite itself.
Additionally, the development of two assay techniques was an integral part of this study. The protein A bacterial adsorbent co-agglutination test was conceived for the quantitation of soluble antigens found in culture supernatant fluids. These antigens were also utilized in the latex agglutination test, a simple and practical serodiagnostic procedure developed for the detection of bovine babesiosis.
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