Sensitive Point-of-Care Detection of Oral Squamous Cell Carcinoma-Associated Salivary miRNA-31 Enabled by Autocycling Primer Extension Reaction and a Bioinspired Photonic-Crystal Microchip

Sensitive, point-of-care detection of salivary miRNA-31 holds considerable potential for the early, noninvasive screening and diagnosis of oral squamous cell carcinoma (OSCC). This study reports the first development of a sensitive, portable biosensor that integrates an autocycling primer extension reaction (ACPER) with a bioinspired photonic-crystal (PC) microchip for quantitative analysis of miRNA-31 in saliva. The high amplification efficiency of ACPER enables sensitive detection of miRNA-31 in saliva. Meanwhile, due to the fluorescence-enhancing properties of the PC microchip, the fluorescence signal generated by the ACPER on the PC microchip can be directly visualized and captured through a smartphone imaging system under ultraviolet-light illumination. The fluorescence intensity values are subsequently extracted via image processing software for quantification analysis. This approach obviates the need for bulky instrumentation and mitigates errors arising from subjective interpretation of color depth by the naked eyes, thereby significantly enhancing detection accuracy. Preliminary clinical feasibility assessment demonstrated that this newly developed biosensor can differentiate between cancer patients and healthy individuals in clinical samples with good accuracy (area under the curve = 1), providing a novel paradigm for the early, noninvasive, and sensitive diagnosis of OSCC.