Rapid solid-phase radioimmunoassay for detection of equine infectious anemia viral antigen and antibodies: parameters involved in standardization.
Solid-phase radioimmunoassays (SPRIA) are described for the detection of equine infectious anemia (EIA) viral antigen and antibodies. Protein-antigen P29 currently used in the agar-gel immunodiffusion (AGID) test was used as antigen in the SPRIA. Rabbit sera selected from positive AGID test data were used to standardize the method. Briefly, wells of flexible microtitre plates coated with antigen were incubated with antiserum followed by a secondary labelled antibody. The radioactivity remaining in the wells after washing provided a measure of the amount of specific antibodies in the serum. When testing a group of rabbit sera, negative for EIA virus antibodies by the AGID test, in the SPRIA a range of positive reactivities was noted. The specificity of the reaction was assessed by inhibition with the antigen. The reaction of immune serum against EIA-virus antigen adsorbed to the wells, was completely inhibited by the antigen in solution. This property was applied in an indirect competitive SPRIA for the detection of viral protein P29. The detection threshold of the SPRIA for EIA virus protein was about 5 ng and about 1 ng of antibody can be detected. The assay is rapid, specific and sensitive and allows the testing of multiple serum samples with the advantage of employing a single secondary labelled antibody.
D000818:Animals; D000914:Antibodies, Viral / Q000032:analysis*; D000956:Antigens, Viral / Q000032:analysis*; D004859:Equine Infectious Anemia / Q000175:diagnosis* / Q000276:immunology; D006736:Horses; D004860:Infectious Anemia Virus, Equine / Q000276:immunology*; D011817:Rabbits; D011863:Radioimmunoassay / Q000379:methods / Q000592:standards; D014764:Viral Proteins / Q000032:analysis / Q000276:immunology
D000914:Antibodies, Viral; D000956:Antigens, Viral; D014764:Viral Proteins