title: Quantification of HIV-RNA from dried blood spots using the Siemens VERSANT® HIV-1 RNA (kPCR) assay.
abstract: OBJECTIVE
  Simplified methods for virological monitoring in resource-limited settings are increasingly needed. We evaluated the performance of the VERSANT(®) 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°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.
authors.affiliation: Department of Therapeutic Research and Medicines Evaluation,  Istituto Superiore di Sanità, Rome,  Italy.
authors.forename: Maria Franca
authors.initials: MF
authors.lastname: Pirillo
authors.forename: Patricia
authors.initials: P
authors.lastname: Recordon-Pinson
authors.forename: Mauro
authors.initials: M
authors.lastname: Andreotti
authors.forename: Maria Grazia
authors.initials: MG
authors.lastname: Mancini
authors.forename: Roberta
authors.initials: R
authors.lastname: Amici
authors.forename: Marina
authors.initials: M
authors.lastname: Giuliano
mesh_terms: D001331:Automation / Q000379:methods; D001769:Blood / Q000821:virology*; D003890:Desiccation / Q000379:methods*; D015658:HIV Infections / Q000821:virology*; D015497:HIV-1 / Q000302:isolation & purification*; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D012367:RNA, Viral / Q000097:blood; D011933:Reagent Kits, Diagnostic; D012680:Sensitivity and Specificity; D013048:Specimen Handling / Q000379:methods*; D019562:Viral Load; D014773:Virology / Q000379:methods
chemical_list: D012367:RNA, Viral; D011933:Reagent Kits, Diagnostic