Development and evaluation of serological screening based on one dried plasma spot for HIV, syphilis, and HCV

Abstract

Background: In the effort to prevent and control HIV/AIDS, China has established a national sentinel surveillance system. However, some sentinel sites face limitations in environmental resources and accessibility, prompting the exploration of alternative sample strategies. Dried plasma spots (DPS) samples are viewed as promising alternatives to traditional plasma samples due to their advantages, including sample stability, easy storage, and convenient transport. This study aims to develop a method for screening HIV, Treponema pallidum (TP), and Hepatitis C Virus (HCV) using DPS samples and assess their performance.

Methods: Based on existing commercial assay kits, a detection method was established through the optimization of experimental parameters, including the amount of plasma on filter paper, the volume of elution solution applied to dried plasma spots, the size of dried plasma spots, elution solution volume, elution solution components, elution temperature, and elution time. A series of laboratory evaluation panels were constructed for laboratory assessments, including the laboratory basic panel, laboratory interference panel, and laboratory precision panel. Additionally, clinical samples were used for evaluation.

Results: Optimal conditions for DPS sample extraction were: plasma volume, 100 µL; DPS size, whole spot; eluent volume, 500 µL; eluent, PBS with 1‰ Tween20; elution time, 2 h; elution temperature, room temperature. A total of 619 paired plasma/DPS samples were tested by both methods. The DPS-based ELISA method exhibited 100% sensitivity/specificity for HIV, 98.6%/100% for TP, and 99.6%/100% for HCV. Kappa values between the plasma samples and DPS samples were 100% for HIV, 99% for TP, and 100% for HCV. The DPS-based ELISA method failed to detect 1 HCV mono-infected sample and TP in 1 HIV/HCV/TP co-infected sample. For the HIV/HCV/TP co-infected sample, the S/CO in the plasma sample was 2.143 and in the DPS sample was 0.5. For HCV, the S/CO (sample OD/cut-off) was 3.049 in the plasma sample and 0.878 in the DPS sample.

Conclusions: A single DPS, following one-time standardized processing, can be used to detect HIV, HCV, and TP. Researching and establishing laboratory testing methods better suited for China's sentinel surveillance have significant practical applications in improving HIV testing in resource-constrained environments.

Keywords: Dried plasma spots; ELISA; Hepatitis C virus; Human immunodeficiency virus; Treponema pallidum.

Fig. 1

Comparison of ELISA tests for detection of anti-HIV/anti-TP/anti-HCV in DPS samples with different volume of plasma. Note: The figures in brackets represent different serial dilution

Fig. 2

Comparison of ELISA tests for detection of anti- HIV/anti-TP/anti-HCV under different combinations based on size of DPS and volume of eluent Notes: A indicates that DPS sample contained 100 µL plasma; B represents the size of DPS sample [whole spot, 1/2 DPS, 1/4 DPS, 2 discs and 1 discs (6 mm)]; C show that the volume of eluent [1 mL, 500 µL, 300 µL,200 µL]; D means the amount of sample according to test kits

Fig. 3

Different elution buffer effects on the DPS-based ELISA method for detection of anti-HIV, anti-TP and anti-HCV

Fig. 4

The variation of test results of anti-HIV, anti-TP and anti-HCV at different elution temperature with the elution time

Fig. 5

Working curves of anti-HIV, anti-TP and anti-HCV in plasma samples and in DPS samples by ELISA. Note: a has been published in Chinese Journal of AIDS & STD and used as a reference in this study