Fortified Donor–Acceptor Spectral Overlap Facilitated Fluorescence Quenching Efficiency for Developing Sensitive Nanometal Surface Energy Transfer-Based Immunochromatographic Test Strips

Constructing a fluorescence quenching immunochromatography test strip (FQ-ICTS) is conducive to sensitive point-of-care testing, yet inefficient quenching efficiency (QE) confines the progress of the FQ-ICTS. Herein an innovative nanometal surface energy transfer (NSET)-based FQ-ICTS with magnificent QE was proposed for the highly sensitive detection of T2 toxin utilizing time-resolved fluorescent microspheres (TRFMs) conjugated with T2-BSA as donors and monoclonal antibodies (mAbs) labeled with spherical and flower-shaped gold particles (AuNPs) as acceptors. As the key to biosensors, by regulating the distinct absorption wavelengths of AuNPs-mAb (520-605 nm), the effect of the donor-acceptor overlapping area on the QE was explored, and the NEST pair with the highest QE was screened out to achieve perfect sensitivity. It was found that the QE increased with the enlargement of the overlapping integral area. At the maximum spectral overlap area (4.68 × 10²³ M^-1 cm^-1 nm⁴), the AuNPs-605 and TRFMs were the optimized NSET pair with the QE of up to 92.7%, and the described FQ-ICTS revealed a limit of detection reaching down to 0.034 ng/mL, which was a 13.2-fold improvement over that of a conventional AuNPs-ICTS, possessing favorable specificity and repeatability. Importantly, the proposed platform could be well-applied in a real maize sample for T2 monitoring with satisfactory recoveries (95.5% to 108.7%), correlating well with the results from HPLC-MS/MS. Concisely, this work demonstrates that the expansion of the donor-acceptor spectral overlap opens up distinct opportunities for the exploration of high QE acceptors and superior performance for an ICTS.