A dual-chamber “one-pot” CRISPR/Cas12a-based portable and self-testing system for rapid HPV diagnostics

Long-lasting infection of high-risk HPV infection has long been recognized as the leading cause of cervical cancer and has garnered widespread attention. However, in resource-limited areas, effective HPV screening is often not easily accessible, resulting in a continuous growth in the incidence and mortality rates of cervical cancer in these areas. In this study, we present a rapid, efficient, and accurate method for HPV detection based on the CRISPR/Cas12a dual-chamber “one-pot” test (DROPT) system. Clinical samples are able to be rapidly tested in the DROPT system to get quantitative fluorescence readout with a simple one-time press. The entire reaction process, including RPA and CRISPR assays, happens within 30 minutes in a portable device composed of an isothermal heating chamber, laser diode source, and silicon photodiode. The results demonstrate that this approach achieves a remarkable limit of detection of 10-18 M. In the testing of HPV16/18 in 60 cervical swab samples, the system reached a sensitivity and specificity of 100% and 95.8%, respectively. Consequently, the DROPT system offers ease of use, requires no large-scale equipment or laboratory personnel, and effectively avoids aerosol contamination, which potentially fills the gap for HPV self-testing and on-site nucleic acid testing.