A sensitive radioimmunoprecipitation assay for human immunodeficiency virus (HIV)

Abstract

A sensitive and efficient radioimmunoprecipitation procedure is described which provides an alternative to Western blotting assays for characterizing antibodies directed against human immunodeficiency viruses (HIV-1). Reaction of solubilized preparations of HTLV-III with 125I-labeled Bolton-Hunter reagent leads to the efficient labeling of all of the major virus-specific proteins, including gp120, gp41, RT (p66/p51), p24, and p17. These labeled proteins are readily immunoprecipitated by immune human sera, by specific sera derived from hyperimmunized animals, and by monoclonal antibodies. This procedure, referred to as BH-RIP, provides a simple assay for characterizing and titering antibodies against HIV which is equivalent in specificity, and more sensitive and efficient than the Western blotting method. In addition, viral proteins labeled in this way are suitable for biochemical studies. In one such application, the number of high-mannose and complex oligosaccharide side chains of gp120 and gp41 were determined by examining the sensitivities of the two viral glycoproteins labeled by this procedure to the glycosidases Endo H and PNGase F.