Identification of immunodominant Trypanosoma musculi antigens recognized by monoclonal antibody and curative immunoglobulin G2a antibody.
Trypanosoma musculi obtained from normal or irradiated (900 rad) hosts or from in vitro cultures were lysed and analysed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). Similar protein banding patterns with a molecular weight (mol. wt) range from 34 to 68 kDa were observed between the two bloodstream forms. In comparison, lysates of cultured parasites showed a unique banding pattern of antigens within the same mol. wt range. Western blot of bloodstream form lysates, probed with immune plasma (IP), revealed a wide range of parasite proteins. However, when probed with the IgG2a-enriched fraction of IP, a major band of approximately 66 kDa was detected on the blot. Several bands of higher mol. wt were also observed. When anti-T. musculi monoclonal antibodies were used to probe the blot, the 66 kDa protein was again recognized. Using indirect fluorescence, live bloodstream form parasites were analysed by flow cytometry and the p66 protein was determined to be a surface molecule. Finally, lysates of 35S-methionine-labelled trypanosomes were immunoprecipitated with Sepharose linked anti-T. musculi monoclonal antibodies and the eluted ligand analysed by SDS-PAGE and autoradiographed. The 66 kDa band was identified, therefore confirming that this protein was of parasite origin.
Shaw
K T
KT
Montreal General Hospital Research Institute, Quebec, Canada.
Shaw
I T
IT
Ryan
P
P
Stevenson
M M
MM
Kongshavn
P A
PA
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000953:Antigens, Protozoan / Q000032:analysis* / Q000276:immunology; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D015151:Immunoblotting; D007074:Immunoglobulin G / Q000276:immunology*; D008297:Male; D051379:Mice; D011233:Precipitin Tests; D014345:Trypanosoma / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000953:Antigens, Protozoan; D007074:Immunoglobulin G