DETECTION OF HIV-1 INFECTION IN-VITRO USING NASBA - AN ISOTHERMAL RNAAMPLIFICATION TECHNIQUE

Establishment of a sensitive infection assay for HIV-1 is essential fo r successful screening of antiviral agents and neutralizing antibodies . In this report, an infection assay is described which measures the e xpression of viral genomic RNA and spliced mRNA intermediates in infec ted cells by an amplification-based technique called NASBA. The extrem e sensitivity of this method permits the detection of viral RNA in per ipheral. blood mononuclear cells (PBMC) within 48 h of infection by a low dose of virus. Similarly, spliced HIV-1 mRNA could be detected wit hin 24 h of infection of CEM cells by HIV-1(IIIB). This NASBA-based in fection assay was shown to titer the neutralization of the HIV-1(IIIB) isolate by serum from an infected human and by a monoclonal antibody to gp120. Furthermore, the inhibitory effects of azidothymidine (AZT) and soluble CD4 on HIV-1(IIIB) infection were quantitated by this assa y. The early detection of virus by NASBA minimizes the contribution of secondary infection, thereby permitting more accurate evaluation of a ntiviral agents and neutralizing antibodies. This assay may be useful for the study of infection of phenotypically distinct HIV-1 isolates, which differ in terms of their replication kinetics.