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bioRxiv · 10.1101/2024.05.23.595500

OPTIMIZATION OF DRIED BLOOD SPOT ASSAYS FOR HEPATITIS B VIRUS SURFACE ANTIBODY QUANTIFICATION

Motshosi, P.·Phinius, B. B.·Jongman, M.·Baruti, K.·Bhebhe, L.·Choga, W. T.·Moyo, S.·Gaseitsiwe, S.·Anderson, M.
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Abstract

Dried blood spot (DBS) cards can be used as an alternative sample collection method to plasma, however, there is no optimized elution protocol for DBS cards specifically for hepatitis B surface antibody (anti-HBs) testing. The study aimed to develop a DBS elution protocol for anti-HBs quantification. Our study sought to determine the ideal phosphate-buffered saline (PBS) buffer volume to use by comparing three PBS volumes (300uL, 450uL, and 500uL), and the optimal time to agitate DBS discs on a plate shaker (1hr, 2hrs, 3hrs, and 4hrs) to yield DBS anti-HBs concentrations that are comparable to corresponding plasma anti-HBs concentrations. The optimal DBS storage temperature (25{degrees}C, -20{degrees}C, and -80{degrees}C) was investigated to determine the ideal longterm storage temperature of the cards. Residual samples were used for optimization (2019-2021). A total of 50 DBS-plasma pairs was used throughout the study, with plasma anti-HBs concentrations being used as the golden standard to compare. The analysis of results was carried out by determining the p-values of the Wilcoxon sign rank. A two-way analysis of variance (ANOVA) was also performed to determine the impact of PBS elution volumes, elution time, and storage temperature on the anti-HBs concentration of DBS samples on STATA Version 15.0. No statistically significant difference between the DBS-plasma anti-HBs pairs was observed when using 450 or 500uL of PBS buffer and when samples were agitated for 3 hours (p=0.594, p=0.499 respectively). The optimal storage temperature for DBS cards was 25{degrees}C because the results showed no statistically significant difference between DBS-plasma anti-HBs titers (p=0.594). The two-way ANOVA analysis showed that elution volumes and time had no statistically significant impact on the DBS anti-HBs concentrations, p=0.948 and p=0.381 respectively. Storage temperature had a statistically significant impact on the DBS anti-HBs concentrations, p=0.002. The optimized DBS elution protocol for anti-HBs quantification will help monitor vaccine efficacy in infants due to the low sample volumes required compared to plasma and also can be used for anti-HBs testing in resource-limited areas around the country.

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Motshosi, P., Phinius, B. B., Jongman, M., Baruti, K., Bhebhe, L., Choga, W. T., Moyo, S., Gaseitsiwe, S., Anderson, M.. 2024-05-26. OPTIMIZATION OF DRIED BLOOD SPOT ASSAYS FOR HEPATITIS B VIRUS SURFACE ANTIBODY QUANTIFICATION. https://doi.org/10.1101/2024.05.23.595500

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