A Stable Dried Tube Specimen for Quality Assurance and Training Programs for HIV Rapid Test for Recent Infection

Abstract

The HIV epidemic is still one of the world's most serious public health challenges, affecting about 38 million people worldwide, especially in sub-Saharan African and Southeast Asian countries. In recent years, tests have been developed to discriminate recent from long-term infection in HIV-infected populations, and these tools can help identify new outbreaks and networks of transmission and target prevention and treatment plans. New rapid tests for recent infection are being deployed in point-of-care settings; however, quality assurance programs need to be implemented to ensure consistency and reliability of the results. We have developed a dried tube specimen (DTS) stabilized with disaccharide trehalose as a quality control reagent for rapid recency testing that can be stored unrefrigerated prior to reconstitution at temperatures up to 37°C for up to 12 weeks. Analysis of 10 trehalose-stabilized DTSs showed that they maintained the same recency classification in all of the samples stored at 4°C and 37°C up to 12 weeks and at 56°C for 2 weeks, while the DTSs prepared without trehalose changed their classification from long-term to recent or recent to negative after storage at 37°C for 12 weeks. Development of DTS quality control reagents will facilitate proficiency and training programs, particularly in settings without cold chain capability in field environments. IMPORTANCE Implementation of stabilized dried tube specimens (DTSs) for quality control and training would facilitate HIV recency programs, especially in point-of-care settings without cold chain availability. This study shows that addition of the disaccharide trehalose to DTSs prior to drying the samples increased stability of the samples across a range of temperatures. This finding provides an affordable way to increase the availability of these key reagents for quality control in resource-constrained settings.

Keywords: HIV rapid test; HIV recency infection; human immunodeficiency virus.

FIG 1

Workflow of DTS preparation. HIV-positive plasma samples were selected, then green food coloring was added (0.1%) to samples, and 20 μL of each was dried in a 2-mL tube, with or without 250 mM trehalose. Samples were stored for up to 12 weeks at different temperatures (4°C, 25°C, 37°C, and 54°C). At each time point (1 day, 2 weeks, 6 weeks, and 12 weeks), one aliquot of each sample was reconstituted in 200 μL PBS-Tween overnight and tested with the Sedia Asanté rapid recency assay kit. Numerical results were obtained with a reader provided by the manufacturer.

FIG 2

Stability of the DTS over time at different temperatures. Each line represents the average control, verification, or LT/R reading percent variation from the baseline through 12 weeks at the noted temperatures. The red lines represent the DTS samples prepared with PBS–0.1% Tween plus 250 mM trehalose, and the blue lines represent samples prepared with PBS–0.1% Tween. There were 10 samples, including 4 long-term, 4 recent, and 2 negative. Error bars represent standard errors of the means. ***, P < 0.001 for PBS–0.1% Tween versus PBS–0.1% Tween plus 250 mM trehalose conditions. Data were compared by ANOVA with Tukey’s posttest.

FIG 3

Shift in Asanté assay classification in DTSs stored at 37°C. The panel of 10 DTSs was prepared with (A) or without (B) trehalose (TRE); reader values were plotted as LT/R line versus VER. Samples that shifted classification after storage can be seen, such as blue symbols from samples classified as R in panel A that migrated to the lower left (negative) quadrant. Diamonds represent baseline values, and circles represent data for any subsequent time point (2, 6, and 12 weeks of DTS storage) at 37°C.

FIG 4

Test strips with and without trehalose. Ten representative test strips each are shown for paired samples stored at 4°C, 25°C, or 37°C for 12 weeks. CTR, control line; VER, positive verification line; LT/R, long-term/recent line.