Screening recombinant baculovirus plaques in situ with antibody probes.
A procedure that allows one to directly detect baculovirus recombinants that express foreign proteins of interest in situ using antibody probes is described. Nitrocellulose replicas of recombinant plaques are probed with specific antibodies, and the antibody antigen complexes are detected using alkaline phosphatase conjugated secondary antibody. This procedure was used to isolate a baculovirus recombinant that expresses Vmw65, the Herpes simplex virus transacting factor that is involved in the induction of viral immediate early gene transcription. Immunoscreening directly on plaque lifts is rapid and reliable and facilitates the testing of different transfer vectors for optimization of foreign gene expression.
Capone
J
J
Department of Biochemistry, McMaster University, Hamilton, Ontario, Canada.
D000911:Antibodies, Monoclonal; D015153:Blotting, Western; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D015696:Gene Products, tat; D007158:Immunologic Techniques; D007304:Insect Viruses / Q000235:genetics*; D011994:Recombinant Proteins / Q000032:analysis*; D018139:Simplexvirus / Q000235:genetics; D014157:Transcription Factors / Q000096:biosynthesis / Q000235:genetics; D010948:Viral Plaque Assay / Q000379:methods; D014764:Viral Proteins / Q000032:analysis*
D000911:Antibodies, Monoclonal; D015696:Gene Products, tat; D011994:Recombinant Proteins; D014157:Transcription Factors; D014764:Viral Proteins