Objective To evaluate the value of supplementary nucleic acid testing by analyzing the virus load (VL) testing results in HIV-1 cases with HIV-1 antibody indeterminate and negative results in western blot (WB) assay, and provide evidence for the further improvement of the HIV testing.

Methods From June to December 2024, HIV-1 antibody indeterminate and negative samples from Beijing Laboratory of the Confirmatory Center were tested for VL, and statistical analysis was conducted in combination with their sample source, screening and testing process, screening value, and WB test strips.

Results A total of 295 samples were collected, including 85 indeterminate samples and 210 negative samples. Among the 85 WB-indeterminate samples, 15 (17.65%) had VL results above the lower limit of detection (positive), of which 14 had VL>1×10⁵ copies/mL, and 1 with VL<1000 copies/mL was from a previously infected individual. Among the 210 antibody-negative samples, 7 (3.33%) were VL-positive, all had VL>1×10⁵ copies/mL. Six WB-indeterminate samples contained env bands (gp41 and gp160/gp120), of which 4 were VL-positive (66.67%). The VL positive rates differed significantly among the samples with different screening detection processes. The mean chemiluminescence screening S/CO ratio of VL-positive samples was higher than that of VL-negative samples. Multivariate binary Logistic regression analysis showed that the likelihood of detecting VL in samples with 20<S/CO ≤ 50 (OR=20.45, 95%CI: 2.54–198.99) was 20.45 times higher than that of samples with S/CO<5, and the likelihood of detecting V in samples with S/CO>50 (OR=93.02, 95%CI: 8.39–999.99) was 93.02 times higher than that of samples with S/CO<5.

Conclusion Supplementary nucleic acid detection for HIV-1 -indeterminate and negative samples in WB assay can improve the detection rate of HIV-infection, enhance the diagnostic capability for HIV infection in early phase, and reduce the social and economic burden of the disease.