Paper-based fluorescent sensor based on molecularly imprinted polymers and upconversion nanoparticles for selective and sensitive D-proline detection in human serum and saliva.

D-Proline has been reported as a potential biomarker for gastric (stomach) cancer, creating a demand for practical and rapid analytical approaches for early diagnosis using simple and non-invasive clinical samples. In this work, a microfluidic paper-based analytical device (µPAD) integrating molecularly imprinted polymer-functionalized upconversion nanoparticles (UCNPs@MIPs) was developed for the selective fluorescence-based detection of D-proline. Owing to near-infrared excitation (980 nm), UCNPs provide background-free fluorescence with high photostability, while MIPs serve as artificial receptors to impart molecular selectivity. The sensing mechanism relies on fluorescence quenching of UCNPs@MIPs induced by D-proline through multiphonon relaxation, resulting in an inverse relationship between fluorescence intensity and analyte concentration. Quantitative analysis was achieved using a homemade IR-laser-controlled light box and smartphone-based image capture, with signal evaluation based on blue-channel intensity. The proposed platform exhibited a linear response over the concentration range of 0.01-0.30 mM, with a limit of detection of 0.016 mM. The method was successfully applied to the determination of spiked D-proline in human saliva and serum samples. Owing to its simplicity, low sample consumption, portability, and adaptability, the proposed sensing strategy shows strong potential for on-site analysis and can be extended to broader applications in chemical analysis, biosensing, and environmental monitoring.