CRISPR analysis based on Pt@MOF dual-modal signal for multichannel fluorescence and visual detection of norovirus.

Abstract

Norovirus is a globally prevalent pathogen that causes acute viral gastroenteritis across all age groups, characterized by its high infectivity and low infectious dose. Consequently, the development of rapid, sensitive, and accurate detection technologies for norovirus presents a significant challenge. In this study, we demonstrate a combination of CRISPR-Cas-based reactions with Pt@MOF-linked immunoassay-like assays. This methodology enables both qualitative analysis and colorimetric readouts of Cas12a-mediated DNA/RNA detection at room temperature, as well as the generation of fluorescent signal readout through base deprotonation-induced Pt@MOF cleavage of a fluorogenic substrate. Furthermore, the integration of RPA amplification with noncanonical PAM-designed CRISPR significantly enhances the sensitivity and flexibility of detection, facilitating the extension of this strategy to other targets. Ultimately, the strategy was validated in spiked food samples with a 100% accuracy rate, consistent with RT-qPCR results. Collectively, this work showcases a viable approach for a dual-functional Pt@MOF-based CRISPR biosensing platform for bioanalysis and a flexible, universal strategy based on noncanonical PAM-designed gRNAs.