Background:
At-home blood collection would enable HIV VL assessment for intensive monitoring without frequent access to a clinic – such as for PrEP programs, perinatal or breastfeeding pediatric monitoring, discordant couples, or participation in ART interruption studies. We present the development and qualification of a scalable, high-throughput PCR method using dried blood collected in Tasso-M50 cartridges.
Methods:
We developed a dried blood assay (‘M50 assay’) with samples from 21 people with HIV (5% female, 67% African American, mean age 51 years, NCT03588715). At clinic visits (mean 20.5), we collected a) capillary blood in two 4-well Tasso-M50 devices and b) matched plasma samples (pVL). Between visits, participants self-collected in two devices that were mailed back. Automated RNA extraction and duplicate RT-qPCR reads with dual LTR/GAG FAM-labeled primers were performed (up to 16 M50 PCR reads per timepoint, sourced from 400 uL blood collection). DNase treatment was used in a subset of samples collected during suppressive ART. Assay performance was tested on separate M50 samples prepared from RNA diluted into whole blood.
Results:
M50 assay PCR reagents/instruments were qualified on plasma samples (94% concordance to reference lab), and M50 VL was comparable between home- and clinic-collected samples. In the separate performance test, the M50 assay had 99.5% specificity and an estimated 95% limit of detection of 150 c/mL if all 16 reads from 2 devices are used. However, 64% of clinic visits with undetected pVL had median M50 read ≥ 200 c/mL, due to cell-associated HIV: M50 VL of these discordant samples correlated with total cellular HIV DNA (r = 0.95) and was stable on suppressive therapy (mean slope –3.1% per week [95% CI –8.4 to +2.3%]). DNase reduced M50 VL while preserving detectability of RNA standards, suggesting that ~28% of the stable M50 background is contributed by DNA and the remainder by RNA.
Conclusions:
We introduce and qualify a new assay for total HIV burden in dried whole blood with an at-home sampling device. Discordant samples with high M50 VL and negative pVL are stable over time and reflect contributions from intracellular viral DNA and RNA. Home sampling of total HIV burden may be able to monitor new infections in prophylaxis-directed studies, assess changes in residual cell-associated virus in cure-directed studies, or detect viral rebound after correcting for the stable on-therapy signal.
