Analytical performance of an automated assay quantifying HIV-1 from dried blood spots

Performance of an automated sample preparation and viral load quantification for HIV-positive dried blood spots (DBS) on the Siemens VERSANT(®) kPCR Molecular System has been previously demonstrated with clinical samples.

Evaluation of the analytical performance of the automated assay using HIV-positive DBS prepared from a dilution series.

Over 300 DBS of HIV-1 nucleic acids from a dilution series in lysed whole blood (322 copies/mL to over 1.6 × 10(7)copies/mL) were spotted onto Whatman 903 cards and analyzed to evaluate analytical performance. Cross contamination was examined with a checkerboard pattern of 82 alternating negative and 1 × 10(6)copies/mL samples.

Analytical sensitivity evaluation with a single 50 μL spot demonstrated a limit of detection (LoD) of 866 copies/mL. Above the LoD, linearity (difference between linearized and observed mean values) was within ± 0.10 log, and accuracy (difference between expected and observed mean values) was within ±0.18log. Imprecision for dilution series levels more than two-fold above the LoD was measured as 20% to 27% CV of quantification. No cross contamination was observed.

The HIV-1 DBS Assay performed similarly to the VERSANT HIV-1 RNA 1.0 Assay (kPCR) in assay linearity, accuracy, and imprecision. The assay was sensitive enough to run single 50μL spots and used an unmodified VERSANT(®) SP Module with only a 30 min incubation prior to automated sample preparation. DBS-specific assay calibrators and controls, expressly formulated for easy frozen storage and identical processing to samples, were employed.