Measuring HIV1 RNA stability in dried blood spot specimens using NucliSENS EasyQHIV1v2.0

Abstract number: P540

van Deursen P., Verhoeven A., de Bie P., de Jong J.

Background: The amount of HIV-1 RNA measured in plasma is one of the key parameters for monitoring anti-viral treatment responses in HIV-1 infected individuals. Accurate viral load measurement depends strongly on sample stability. In remote areas, sample collection sites can be located far from test sites, meaning that sample stability and thereby accurate measurement of the clinical state of the treated individual is at risk. To circumvent this possible risk, dried blood spot (DBS) testing is proposed as alternative for plasma testing.

Objectives: The aim of this study is to determine HIV-1 RNA stability in DBS using different storage conditions.

Methods: A total of 5, 10, 4 and 5 EDTA whole blood samples obtained from HIV-1 RNA negative individuals were spiked with HIV-1 RNA: 0 VQA cps/ml, 700 VQA cps/ml, 21,000 VQA cps/ml and 70,000 VQA cps/ml, respectively. For these samples, 50 ml spots were made on paper (ProteinsaverTM 903^® Card, Whatman) and dried 3–48 hours. Next samples were stored at different temperatures (-20°C, 5°C, RT, 37°C, and 55°C). Samples stored at 37°C were subjected first to shipment simulation; 5 days 37°C (of which 8 hours high humidity at 37°C and 8 hours high humidity at 55°C), 3 days 5°C, 5 days -20°C, 3 days 5°C, and subsequently 5 days 37°C (of which 8 hours high humidity at 37°C and 8 hours high humidity at 55°C). High humidity conditions were tested at 37°C and 55°C. After storage HIV-1 RNA levels were measured using two spots (0.1 ml blood) and NucliSENS EasyQ HIV-1 v2.0.

Results: For the high input samples no or limited reduction in HIV-1 RNA levels (0.30 log10) was observed for all conditions tested with one exception (6 weeks 37°C at high humidity). For the low input sample (70 VQA copies/extraction) the detection rate was 80% for all conditions tested, with an overall detection rate of 244/250 (97.6%). Sample stability was demonstrated for a period of 9 weeks at 37°C (with and without including shipment simulation), 3 weeks high humidity at 37°C, 1 week high humidity at 55°C, 3 weeks 4°C, 1 week -20°C, and 5 months room temperature.

Conclusion: HIV-1 RNA stability in DBS was demonstrated for several conditions including 5 months room temperature, shipment simulation, and storage at 37°C for 9 weeks. Instability was observed after 6 weeks 37°C high humidity. The results support the use of DBS specimen for accurate viral load measurements after transport and storage when taken these limitations into account.