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About: Background: Current HIV-1 viral-load assays are too expensive for resource-limited settings. In some countries, monitoring of antiretroviral therapy is now more expensive than treatment itself. In addition, some commercial assays have shown shortcomings in quantifying rare genotypes. Methods: We evaluated real-time reverse transcription-PCR with internal control targeting the conserved long terminal repeat (LTR) domain of HIV-1 on reference panels and patient samples from Brazil (n = 1186), South Africa (n = 130), India (n = 44), and Germany (n = 127). Results: The detection limit was 31.9 IU of HIV-1 RNA/mL of plasma (>95% probability of detection, Probit analysis). The internal control showed inhibition in 3.7% of samples (95% confidence interval, 2.32%–5.9%; n = 454; 40 different runs). Comparative qualitative testing yielded the following: Roche Amplicor vs LTR assay (n = 431 samples), 51.7% vs 65% positives; Amplicor Ultrasensitive vs LTR (n = 133), 81.2% vs 82.7%; BioMerieux NucliSens HIV-1 QT (n = 453), 60.5% vs 65.1%; Bayer Versant 3.0 (n = 433), 57.7% vs 55.4%; total (n = 1450), 59.0% vs 63.8% positives. Intra-/interassay variability at medium and near-negative concentrations was 18%–51%. The quantification range was 50–10 000 000 IU/mL. Viral loads for subtypes A–D, F–J, AE, and AG yielded mean differences of 0.31 log(10) compared with Amplicor in the 10(3)–10(4) IU/mL range. HIV-1 N and O were not detected by Amplicor, but yielded up to 180 180.00 IU/mL in the LTR assay. Viral loads in stored samples from all countries, compared with Amplicor, NucliSens, or Versant, yielded regression line slopes (SD) of 0.9 (0.13) (P <0.001 for all). Conclusions: This method offers all features of commercial assays and covers all relevant genotypes. It could allow general monitoring of antiretroviral therapy in resource-limited settings.

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