Electricity-Free Amplification and Detection for Molecular Point-of-Care Diagnosis of HIV-1

Abstract

In resource-limited settings, the lack of decentralized molecular diagnostic testing and sparse access to centralized medical facilities can present a critical barrier to timely diagnosis, treatment, and subsequent control and elimination of infectious diseases. Isothermal nucleic acid amplification methods, including reverse transcription loop-mediated isothermal amplification (RT-LAMP), are well-suited for decentralized point-of-care molecular testing in minimal infrastructure laboratories since they significantly reduce the complexity of equipment and power requirements. Despite reduced complexity, however, there is still a need for a constant heat source to enable isothermal nucleic acid amplification. This requirement poses significant challenges for laboratories in developing countries where electricity is often unreliable or unavailable. To address this need, we previously developed a low-cost, electricity-free heater using an exothermic reaction thermally coupled with a phase change material. This heater achieved acceptable performance, but exhibited considerable variability. Furthermore, as an enabling technology, the heater was an incomplete diagnostic solution. Here we describe a more precise, affordable, and robust heater design with thermal standard deviation <0.5°C at operating temperature, a cost of approximately US$.06 per test for heater reaction materials, and an ambient temperature operating range from 16°C to 30°C. We also pair the heater with nucleic acid lateral flow (NALF)-detection for a visual readout. To further illustrate the utility of the electricity-free heater and NALF-detection platform, we demonstrate sensitive and repeatable detection of HIV-1 with a ß-actin positive internal amplification control from processed sample to result in less than 80 minutes. Together, these elements are building blocks for an electricity-free platform capable of isothermal amplification and detection of a variety of pathogens.

HIV microarray for the mapping and characterization of HIV-specific antibody responses

“…32,33 Another focus in diagnostic testing is the discrimination of HIV-1 and HIV-2 infections 33 and the availability of tests that can be used for point of care testing and in countries under difficult conditions where laboratory facilities are lacking and costs are an important issue. 34,35 Fast and comprehensive immunoassays such as the HIV microarray may therefore address another increasing need of the scientific HIV-research community. Multiplex tests will be useful for the screening of large numbers of samples in prevalence, population studies and vaccine trials.…”

Expanding the MDx toolbox for filarial diagnosis and surveillance

“…These include electricity-free amplification [65,66] using pocket warmers [67], and new pH-sensitive dyes for improved visual detection [54]. In the future, a lateral flow dipstick or ELISA based on hybridization of LAMP products to specific immobilized probes and detection of the captured amplicons by immunoassay technology is feasible [68].…”

Single-use paper-based hydrogen fuel cells for point-of-care diagnostic applications

“…The duration of the power delivery can be extended as these devices use up to 20x more MgFe for a single test (and consequently, produce as much hydrogen) than the amount used in the fuel cells presented here. [39][40][41][42] This approach implies reusability of the fuel cells for more than one measurement, thus reducing the overall cost per test.…”

“…the corrosion of magnesium iron particles. [36][37][38][39][40][41][42] This chemistry is very convenient, as it , requires only the addition of water to achieve 100°C (a reason why it has also been widely used to heat portable meals). During the course of the chemical reaction the magnesium is oxidized, releasing hydrogen gas as a byproduct, which previously was simply vented off to the atmosphere.…”