Abstract Background Monitoring anti-retroviral therapy requires that viral load assays for human immunodeficiency virus type 1 (HIV-1) be applicable to diverse HIV-1 subtypes. Objectives To evaluate NucliSens EasyQ™ HIV-1 assay for quantitation of common HIV-1 subtypes prevalent in South-east Asia. Study design One hundred and nineteen plasma samples collected in Hong Kong and Cambodia were used to compare the performance of NucliSens EasyQ™ HIV-1 and COBAS Amplicor™ HIV-1 Monitor version 1.5 assays. Viral RNA extracted from the NucliSens MiniMAG™ was also used for HIV-1 subtyping. Results Performance of NucliSens EasyQ™ correlated well with COBAS Amplicor™ ( r = 0.777, p < 0.001) and the small mean difference (0.0462 log 10 IU/mL) obtained in the Bland and Altman model indicated good agreement between two assays. The NucliSens EasyQ™ assay demonstrated a 95% sensitivity at 500 IU/mL and 100% specificity. Reproducibility of this assay was within log 10 2–4 IU/mL and had a coefficient of variation between 2.3% and 10.4%. Among the 109 specimens included in the analysis, HIV-1 subtyping identified 64 CRF01_AE, 38 subtype B, 3 subtype C, 3 CRF07_BC and 1 subtype G viruses. Conclusions Performance of NucliSens EasyQ™ was comparable to COBAS Amplicor™ for HIV-1 viral load monitoring. RNA extracts from NucliSens MiniMAG™ could be used for HIV-1 viral load monitoring, subtyping and drug resistance mutations detection. Our findings highlight the versatility of both NucliSens EasyQ™ and COBAS Amplicor™ in monitoring prevalent subtypes and rare circulating recombinant forms (CRFs) in the South-east Asia region.