CRISPR/Cas13a‐Enhanced Porous Hydrogel Encapsulated Photonic Barcodes for Multiplexed Detection of Virus

Abstract In this study, we present an ultrasensitive and specific multiplexed detection method for SARS‐CoV‐2 and influenza (Flu) utilizing CRISPR/Cas13a technology combined with a hydrogel‐encapsulated photonic crystal (PhC) barcode integrated with hybridization chain reaction (HCR). The barcodes, characterized by core‐shell structures, are fabricated through partial replication of periodically ordered hexagonally close‐packed silicon dioxide beads. Consequently, the opal hydrogel shell of these barcodes features abundant interconnected pores that provide a substantial surface area for probe immobilization. Furthermore, the inherent structural colors remain stable during detection events due to the robust mechanical strength of the barcode cores. This integration of CRISPR/Cas13a and HCR leverages both the highly specific RNA recognition capabilities and trans‐cleavage activity of Cas13a while employing HCR to enhance sensitivity. Upon encountering target RNA, Cas13a cleaves a hairpin probe, thereby initiating subsequent HCR amplification for enhanced detection sensitivity. Our method demonstrates high accuracy and sensitivity in multiplexed detection of SARS‐CoV‐2, Flu A and Flu B RNA with a limit‐of‐detection as low as 200 aM. Importantly, this assay also exhibits acceptable accuracy in repeated clinical sample testing. Thus, our platform represents a promising strategy for highly sensitive multiplexed virus detection in clinical.