Abstract: Objective To developan easy and reliable multinested RT-PCR assay for diagnosis of HIV-1 infection.Methods Several sets of PCR prmiers were designed to develop a multine sted RT-PCR assay for amplification of gag,pol,and envgene of HIV-1 variants prevailing in Guangxi.To tally 118 HIV positive samples and 48 HIV negative samples were detected with the multi nested RT-PCR assay and the detection resultswere compared to that of Western blot.Results The lowe stlmiit of the multine sted RT-PCR a ssay was 250 copies/mL and the sensitivity,specificity,and repeatability was 96.6%,97.9%,and 98.3%,respectively.The consistency bewteen the multine sted RT PCRassay and We stern blot a ssay was 97.6%.Conclusion A multinested RT-PCR assay with high sensitivity,specificity,repeatability,and time saving was developed and could be applied for auxiliary diagnosis of HIV1 infection.