A CRISPR-RCA Enhanced Ru-doped Nanophosphor for Cd2+ Detection in Seafood

Seafood products play a vital role in human dietary. Developing highly sensitive and rapid-response detection strategies is crucial for ensuring the seafood safety and safeguarding public health. In this study, I developed a novel signal-amplified, in-situ, and highly sensitive fluorescence detection platform based on rolling circle amplification (RCA)-assisted CRISPR technology for the efficient detection of cadmium ions (Cd2+) in raw shrimp seafood. Ru(bpy)32+ doped SiO2 (RuSi) nanoparticles are employed as fluorescent probes in conjunction with reporting probes. In the presence of Cd2+, aptamers bind to Cd2+, thereby releasing the primer probe. Stimulated by the primer, the PC3 probe interacts with Phi29 enzyme to induce highly efficient RCA, significantly increasing the number of PC3 fragments in the system. These generated fragments then associate with NTS probes, Cas12, and CrRNA to form CRISPR complexes capable of non-specific single-strand cleavage. These complexes cleave the complementary probe BP of RP in the system, preventing the formation of double-stranded structures between RuSi QDs-RP complexes and BP. Consequently, RuSi QDs form single-strand-graphene complexes with reduced graphene oxide on the substrate surface, generating fluorescence. Under optimized conditions, the sensor exhibits a good linear response to Cd2+ with a detection sensitivity of 1 nM. Furthermore, using this detection system, I designed an integrated fluorescence detector for sample injection, detection, and signal readout. The sensor, powered by an STM32 chip, recognizes fluorescence signals and monitors Cd2+ concentrations in real-time through an LCD display. This sensing device offers advantages such as instant concentration display, portability, and high sensitivity, holding promising prospects for application in the field of food safety detection.