Single-use, electricity-free amplification device for detection of HIV-1

Abstract

Early and accurate diagnosis of HIV is key for the reduction of transmission and initiation of patient care. The availability of a rapid nucleic acid test (NAT) for use at the point-of-care (POC) will fill a gap in HIV diagnostics, improving the diagnosis of acute infection and HIV in infants born to infected mothers. In this study, we evaluated the performance of non-instrumented nucleic acid amplification, single-use disposable (NINA-SUD) devices for the detection of HIV-1 in whole blood using reverse-transcription, loop-mediated isothermal amplification (RT-LAMP) with lyophilized reagents. The NINA-SUD heating device harnesses the heat from an exothermic chemical reaction initiated by the addition of saline to magnesium iron powder. Reproducibility was demonstrated between NINA-SUD units and comparable, if not superior, performance for detecting clinical specimens was observed as compared to the thermal cycler. The stability of the lyophilized HIV-1 RT-LAMP reagents was also demonstrated following storage at -20, 4, 25, and 30°C for up to one month. The single-use, disposable NAT minimizes hands-on time and has the potential to facilitate HIV-1 testing in resource-limited settings or at the POC.

Keywords: Diagnosis; HIV-1; Nucleic acid amplification; Point-of-care; RNA.

Figure 1. NINA-SUD device

(A) A cross-sectional diagram of the NINA-SUD device. Removable foam insulation lid (a) allows insertion of 3– 100uL PCR tubes (b) into the device. An exothermic chemical reaction fuel pouch (d) delivers heat to palmitic acid PCM (c), buffering temperature to amplification specification. The reaction by-product steam is channeled into a condensing chamber (f). The PCR tube holder and support structure is made from rapid-prototyped plastic (e). (B) A photograph of the NINA-SUD device before and after initiation of the exothermic chemical reaction, along with the PCR tubes post-reaction. From left to right, the PCR tubes contain a sample, positive control, and negative control.

Figure 2. NINA-SUD temperature profile

The average temperature of 10 NINA-SUD test devices (black line), recorded for 60 minutes post-initiation of chemical reactants. The shaded gray area of the plot represents the standard deviation.

Figure 3. Lyophilized RT-LAMP reagent stability

The total amplification time until positivity is plotted for DNA (A) and RNA (B) over days the lyophilized reagents were stored at temperatures of −20°C (solid line, closed circle), 4°C (solid line, open circle), 25°C (dashed line, closed circle), and 30°C (dashed line, open circle).