A simple method to elute cell-free HIV from dried blood spots improves their usefulness for monitoring therapy.

Background: Dried blood spots (DBS) improve access to HIV viral load (VL) testing, but yield increased VL measurements compared to the plasma reference method because of cell-associated viral nucleic acid. In clinical settings, DBS methods may falsely categorize many patients as failing therapy.

Objectives: Description of a simple method, free virus elution (FVE), to preferentially elute plasma-associated virus from DBS samples with phosphate-buffered saline, and an initial HIV VL performance comparison with standard DBS elution methods.

Study Design: The mechanism of action of FVE was studied with model DBS samples containing purified virus or washed HIV-containing cells, and with a DNA-specific HIV PCR. Using clinical samples, VL results from the new FVE method were compared to results from a dried fluid spot procedure (DFSP) protocol, which uses a guanidinium-based elution method, using plasma VL as the reference method.

Results: Model system experiments suggest that the method efficiently separates virus from cell-associated HIV, with a wide tolerance for incubation time and temperature. In 196 clinical samples, FVE reduced VL over-quantification from DBS, and improved DBS clinical concordance with plasma from 67% to 95%.

Conclusions: A simple elution in PBS significantly reduced the over-quantification of HIV VL in DBS. Additional studies are needed to validate the method in fingerstick-collected specimens and to further understand the compartmentalization of HIV DNA and RNA in DBS specimens.