Near-infrared light-controlled catalytic hairpin assembly fluorescent biosensor for the sensitive and specific detection of fibroblast activation protein

Fibroblast activation protein (FAP) is significantly upregulated in activated cardiac fibroblasts after myocardial infarction, and its serum concentration is negatively correlated with cardiac function, infarct size, and ischemia score. It is an ideal biomarker for early diagnosis and prognosis evaluation of cardiovascular disease. In this work, a fluorescent biosensor based on a near-infrared light-controlled catalytic hairpin assembly (CHA) signal amplification strategy was developed for highly sensitive and specific detection of FAP. First, upconversion nanoparticles (UCNPs) are synthesized, and then the surface of the UCNP is modified with sodium citrate via electrostatic adsorption to introduce carboxyl groups, resulting in the formation of carboxyl-functionalized UCNPs. Finally, they were covalently coupled with the photo-controlled hairpin probe (H1) through amide condensation. Under 808 nm illumination, UCNPs emit ultraviolet light to cleave the photocleavable (PC) linker of H1, exposing the S1 binding site. Competitive displacement of free S1 by target FAP initiates CHA cycle amplification, achieving significant recovery of Cy5 fluorescence. Under optimal conditions, the biosensor exhibited a good linear response to FAP, with a detection range of 0.5-6 ng mL^-1 and a limit of detection as low as 0.384 ng mL^-1. The results of the human serum spiked recovery experiment revealed a recovery rate of 98.2% to 103.4%, with an RSD of less than 4.6%. The constructed biosensor has the benefits of high sensitivity, specificity, good stability, and resistance to matrix interference, demonstrating good potential for application in FAP-related detection.