Moving toward rapid and low-cost point-of-care molecular diagnostics with a repurposed 3D printer and RPA

Traditionally, the majority of nucleic acid amplification-based molecular diagnostic tests are done in centralized settings. In recent years, point-of-care tests have been developed for use in low-resource settings away from central laboratories. While most experts agree that point-of-care molecular tests are greatly needed, their availability as cost-effective and easy-to-operate tests remains an unmet goal. In this article, we discuss our efforts to develop a recombinase polymerase amplification reaction-based test that will meet these criteria. First, we describe our efforts in repurposing a low-cost 3D printer as a platform that can carry out medium-throughput, rapid, and high-performing nucleic acid extraction. Next, we address how these purified templates can be rapidly amplified and analyzed using the 3D printer's heated bed or the deconstructed, low-cost thermal cycler we have developed. In both approaches, real-time isothermal amplification and detection of template DNA or RNA can be accomplished using a low-cost portable detector or smartphone camera. Last, we demonstrate the capability of our technologies using foodborne pathogens and the Zika virus. Our low-cost approach does not employ complicated and high-cost components, making it suitable for resource-limited settings. When integrated and commercialized, it will offer simple sample-to-answer molecular diagnostics.