Rapid detection of HPV16 utilizing recombinase polymerase amplification with the employment of an extremely low concentration of the probe†

Human papillomavirus 16 (HPV16) infection is the leading cause of cervical cancer. The current mainstream method for detecting HPV16 is quantitative real-time PCR (qPCR). However, due to its time-consuming nature and reliance on expensive qPCR instruments, there is growing interest in more convenient, rapid and private approaches such as recombinase polymerase amplification (RPA). In this study, we innovated an effective method for HPV16 detection by integrating a RPA system with lateral flow strip (LFS) detection. Primers with optimal efficiency and specificity were designed, and false positive signals caused by dimers were eliminated by reducing the probe concentration. The RPA-LFS method demonstrates high sensitivity and specificity, capable of detecting 230 copies per reaction of HPV16 within 25 min without cross-reactivity to other subtypes. It exhibits good tolerance, remaining unaffected by 1.0% miconazole, 0.5% tioconazole and 1.0% hemoglobin. The results of clinical samples detected by this method were consistent with those of qPCR. The method provides a practical reference for HPV16 diagnosis and can be valuable in home and resource-limited settings, contributing to the reduction of cervical cancer incidence.