Advances and challenges of fully integrated paper-based point-of-care nucleic acid testing

Nucleic acid testing (NAT) has been studied extensively in paper-based diagnostics, presenting a substantially higher sensitivity and specificity than immunoassays. Paper-based NAT provides an alternative to laborious, expensive and time-consuming conventional NAT. Recent advances in paper fabrication and modification technologies have made it possible to integrate all key steps of NAT (i.e., sample preparation, nucleic acid amplification and amplicon detection) into one single paper-based device, and are hence suitable for resource-poor settings. However, multiple challenges are yet to be addressed to translate the technologies into practical applications. In the present review, we discuss the current status and challenges in accomplishing each key step of NAT using low-cost paper substrates. We highlight the most recent advances, challenges and possible solutions in integrating all these steps into a compact paper-based device. We also review the latest progress towards commercialization and future perspectives on the development of an ideal sample-in-answer-out device.