Development of a Microfluidic Point-of-Care Platform for HPV Detection Based on Helicase-Dependent Amplification

Human papillomavirus (HPV) is the most prevalent sexually transmitted infection worldwide and a leading cause of cervical cancer, accounting for over 300, 000 deaths annually, primarily due to high-risk genotypes HPV-16 and HPV-18. Conventional molecular diagnostic methods, such as polymerase chain reaction (PCR), require expensive instrumentation and well-equipped laboratories, which limits their applicability in low-resource or decentralized settings. To address this challenge, the aim of this study was to develop a prototype point-of-care (POC) diagnostic platform based on helicase-dependent amplification (HDA) integrated into a microfluidic device for the specific detection of HPV-16 and HPV-18. The proposed POC platform comprises a disposable poly (methyl methacrylate) (PMMA) microfluidic device, a portable warming mat for isothermal amplification at 65 °C, and a compact electrophoresis chamber for fluorescence-based visualization using SYBR Safe dye, with an approximate total cost of 320 USD. Platform validation was performed on 33 samples, demonstrating amplification of target sequences in less than 60 min with only 20 µL of reaction volume, a limit of detection (LOD) of 15 copies (cp) per reaction, a sensitivity of 95. 52%, and a specificity of 100%. This portable and scalable platform constitutes a cost-effective and reliable tool for the detection of HPV, supporting global health initiatives, including those driven by the World Health Organization (WHO), aimed at eliminating cervical cancer as a public health threat, as it can be implemented in decentralized or resource-limited settings.