Quantification of HIV-RNA from dried blood spots using the Siemens VERSANT® HIV-1 RNA (kPCR) assay

Objectives: Simplified methods for virological monitoring in resource-limited settings are increasingly needed. We evaluated the performance of the VERSANT (R) HIV-1 RNA (kPCR) assay for the determination of HIV-1 viral load from dried blood spots (DBS). Assay sensitivity and correlation with plasma quantification values were assessed. Methods: A total of 98 DBS were prepared from fresh blood samples of HIV-infected patients. DBS were kept at room temperature for 6 weeks or 7 months before processing while the corresponding plasma samples were stored at -80 degrees C. DBS were first pre-treated in a special DBS buffer. The DBS extracts and the plasma samples were then purified and amplified using the VERSANT assay reagents. Results: In the first series of tests, performed after 6 weeks of storage, there was good correlation between quantification of viral load in plasma and in DBS (r=0.95, P<0.001). The detection rate in DBS was 100% when plasma levels were >1000 copies/mL. The sensitivity and specificity of the DBS assay were 88.2% [95% confidence interval (CI) 79.4-93.6] and 69.2% (95% CI 42.0-87.4), respectively. Using the 5000 copies/mL threshold (defining virological failure in resource-limited settings), both positive and negative predictive values were high (95.2% and 87.5%, respectively). After 7 months of storage there was a modest decrease in the detection rate and less significant correlations for samples with HIV-RNA <5000 copies/mL. Conclusions: Quantification of HIV-RNA from DBS by the VERSANT automated sample preparation and detection method can be used to diagnose virological failure in HIV-positive patients.