Sensitive and Quantitative Point-of-Care HIV Viral Load Quantification from Blood Using a Power-Free Plasma Separation and Portable Magnetofluidic Polymerase Chain Reaction Instrument

Point-of-care (POC) HIV viral load (VL) tests are needed to enhance access to HIV VL testing in low- and middleincome countries (LMICs) and to enable HIV VL self-testing at home, which in turn have the potential to enhance the global management of the disease. While methods based on real-time reverse transcription-polymerase chain reaction (RT-PCR) are highly sensitive and quantitatively accurate, they often require bulky and expensive instruments, making applications at the POC challenging. On the other hand, although methods based on isothermal amplification techniques could be performed using lowcost instruments, they have shown limited quantitative accuracies, i.e., being only semiquantitative. Herein, we present a sensitive and quantitative POC HIV VL quantification method from blood that can be performed using a small power-free three-dimensionalprinted plasma separation device and a portable, low-cost magnetofluidic real-time RT-PCR instrument. The plasma separation device, which is composed of a plasma separation membrane and an absorbent material, demonstrated 96% plasma separation efficiency per 100 mu L of whole blood. The plasma solution was then processed in a magnetofluidic cartridge for automated HIV RNA extraction and quantification using the portable instrument, which completed 50 cycles of PCR in 15 min. Using the method, we achieved a limit of detection of 500 HIV RNA copies/mL, which is below the World Health Organization's virological failure threshold, and a good quantitative accuracy. The method has the potential for sensitive and quantitative HIV VL testing at the POC and at home self-testing.