Bacterial expression of the acquired immunodeficiency syndrome retrovirus p24 gag protein and its use as a diagnostic reagent.

Abstract

A retrovirus [lymphoadenopathy-associated virus, human T-cell leukemia virus type III, acquired immunodeficiency syndrome (AIDS)-related virus] suspected of causing AIDS has been isolated recently. The detection of exposure to this retrovirus in donors of various blood products is important to prevent transmission of the disease from these donors to recipients. In the majority of cases, the detection of antibodies directed against either the viral core protein, a Mr approximately equal to 24,000 protein termed p24 gag, or the viral envelope antigen is proof of previous viral infection. Thus, we have expressed the p24 gag antigen in Escherichia coli in order to produce a diagnostic reagent for the detection of virus exposure. The bacterially synthesized antigen reacts with human and rabbit antisera directed against the native p24 gag protein in both electrophoretic transfer blot assay and ELISA. In addition, the use of bacterially produced antigens for ELISAs gave results that were comparable to those obtained by using antigens isolated from the virus.

Sensitivity and Specificity of a Nested Polymerase Chain Reaction for Detection of Lentivirus Infection in Lions (Panthera leo)

“…The FIV ELISA uses recombinant gag proteins that are believed to be highly immunogenic, yet conserved among feline lentiviruses. 9,12 Overall, there was 83% agreement (20/24) between the two forms of antibody testing, with only four discrepant results (Table 1). All four of the discrepant results were positive when using the FIV ELISA and negative when using the FIVpco ELISA.…”

The purification of eukaryotic polypeptides synthesized in Escherichia coli

“…Receptor binding of human complement fragment C5a (Mandecki et al, 1985) and biological activity of FMDV proteinase (Klump et al, 1984) were demonstrated using unpurified lysis supernatants. Pure retroviral p24 gag protein was purified from E. coli in a single immunopurification step (Dowbenko et al, 1985) while a five-step process was used to produce human growth hormone (Olsen et al, 1981). Other examples of soluble proteins are human IFN-a (Staehelin et al, 1981;De Maeyer et al, 1982) boVine IFN-a (Grosfeld et al, 1985), chicken triosephosphate isomerase (Straus & Gilbert, 1985), human lymphotoxin (Gray et al, 1984), human TNF (Pennica et al, 1984) and murine TNF .…”

Seven Human Immunodeficiency Virus (HIV) Antigen-Antibody Combination Assays: Evaluation of HIV Seroconversion Sensitivity and Subtype Detection

“…These assays used the solid phase coated with viral antigens and polyclonal antibodies to human immunoglobulins conjugated to an enzyme for detection of HIV-specific antibodies (1,24). The second-generation assays used HIV recombinant antigens instead of viral lysate as the source of antigen on the solid phase and also incorporated recombinant antigen for HIV-2 (6,10,11,12,13,16). These assays had improved specificity though the overall sensitivity remained similar to that of the first-generation assays.…”