Bimodal persistent luminescence for autofluorescence-free ratiometric biosensing

In optical biosensing, analyte-independent factors such as autofluorescence interference and excitation source fluctuation decrease the sensitivity and accuracy. Herein, we reported a bimodal persistent luminescence strategy to design dual-emissive persistent luminescence nanoparticles (PLNPs) with built-in self-calibration to preclude interference from analyte-independent factors in biosensing. As a proof of concept, ZnGa₂O₄:Cr PLNPs with emissions at both 490 nm and 695 nm were designed. The I₄₉₀/I₆₉₅ ratio of ZnGa₂O₄:Cr was readily adjusted by simply changing the doping concentration of Cr³⁺. The ZnGa₂O₄:Cr PLNPs were employed for the ratiometric detection of urinary mesna. A good linear relationship between the I₄₉₀/I₆₉₅ ratio of ZnGa₂O₄:Cr-based nanoprobe and the concentration of mesna was obtained in the range of 0-40 μM. The limit of detection was about 0.40 μM. Results showed that autofluorescence interference from urine was totally eliminated by collecting the persistent luminescence signal of ZnGa₂O₄:Cr after excitation ceased. Moreover, the built-in self-calibration feature of the ratiometric ZnGa₂O₄:Cr PLNPs efficiently suppressed the interference from fluctuations in instrumental parameters during urinary mesna detection. The recovery rates of mesna in the spiked urine samples are in the range of 99.1~109.0%, showing the reliability of the ratiometric ZnGa₂O₄:Cr PLNPs in urinary mesna detection. ZnGa₂O₄:Cr can further be expanded to the detection of other analytes in complex matrices. This study may open new opportunities for the design of dual-emissive PLNPs with tunable ratios of emission intensity, and it can further promote the applications of optical biosensing in disease diagnosis, food safety, and environmental monitoring.