An immunoaffinity purification procedure for SV40 large T antigen.
A rapid purification procedure for SV40 large T antigen has been developed which combines the use of an adenovirus-SV40 hybrid virus which overproduces large T antigen, and immunoaffinity chromatography on an anti-large T monoclonal antibody coupled to protein A Sepharose. The protein exhibits the p53-binding, ATPase, and sequence-specific DNA-binding activities of T antigen. The purification procedure can be completed in 1 day and allows the isolation of milligram amounts of large T in excellent yield. The pure protein is extremely antigenic and is tolerant of iodination to high specific activity, permitting the development of a competition radioimmunoassay for large T that reliably detects nanogram amounts of the protein.
D000818:Animals; D000936:Antigen-Antibody Complex; D000952:Antigens, Polyomavirus Transforming; D000957:Antigens, Viral, Tumor / Q000235:genetics / Q000302:isolation & purification*; D001483:Base Sequence; D002460:Cell Line; D002522:Chlorocebus aethiops; D002846:Chromatography, Affinity / Q000379:methods; D004268:DNA-Binding Proteins; D004587:Electrophoresis, Agar Gel; D007668:Kidney; D009856:Oncogene Proteins, Viral / Q000235:genetics / Q000302:isolation & purification*; D011863:Radioimmunoassay; D013539:Simian virus 40 / Q000235:genetics / Q000276:immunology*
D000936:Antigen-Antibody Complex; D000952:Antigens, Polyomavirus Transforming; D000957:Antigens, Viral, Tumor; D004268:DNA-Binding Proteins; D009856:Oncogene Proteins, Viral