Afterglow Resonance Energy Transfer Inhibition for Fibroblast Activation Protein-α Assay

Traditional photoluminescence resonance energy transfer (PRET)-based sensors are widely applied, but still suffer from the severe background interference from in situ excitation. The afterglow nature of the persistent luminescence nanoparticles (PLNPs) allows optosensing after the stoppage of in situ illumination, and thus subtly overcomes that interference. We proposed a simple strategy for functionalizing PLNPs for bioanalytical applications and the new afterglow resonance energy transfer (ARET)-based assay for quantitative determination and imaging of fibroblast activation protein-alpha (FAPα) in live cells using Au-decorated Cr³⁺_0.004:ZnGa₂O₄ as donor and Cy5.5-KGPNQC-SH as acceptor. The ARET between the donor and acceptor quenches the afterglow of the donor, and the cleavage of peptide KGPNQC by FAPα inhibits the ARET and restores the afterglow of the donor. The ARET-based assay of FAPα, with the linear range of 0.1-2.0 mg·L^-1 (1.2-22.9 nM), LOD of 11 μg·L^-1 (115 pM), and RSD of 3.9% (for 0.5 mg·L^-1 FAPα, n = 5), displays higher sensitivity, lower limit of detection (LOD), and better anti-interference capability than the corresponding PRET-based assay. Besides, the ARET-based sensors are lighted up by the FAPα-positive U87MG and MDA-MB-435 cells, but kept in the dark when incubated in the FAPα-negative AD293 cells. The proposed ARET-based sensor can detect FAPα of U87MG and MDA-MB-435 living cells in human serum with the spiked recoveries of 95.6-103%. Our data demonstrated a simple and effective strategy for bridging PLNPs to bioanalytical applications, and an attractive ARET assay of FAPα.