A universal CRISPR/Cas12a-powered intelligent point-of-care testing platform for multiple small molecules in the healthcare, environment, and food.

Growing studies focusing on nuclear acid detection via the emerging CRISPR technique demonstrate its promising application. However, limited works solve the identification of non-nucleic acid targets, especially multiple small molecules. To address challenges for point-of-care testing (POCT) in complex matrices for healthcare, environment, and food safety, we developed CRISPR Cas12a-powered highly sensitive, high throughput, intelligent POCT (iPOCT) for multiple small molecules based on a smartphone-controlled reader. As a proof of concept, aflatoxin B1 (AFB1), benzo[a]pyrene (BaP), and capsaicin (CAP) were chosen as multiple targets. First, three antigens were preloaded in independent microwells. Then, the antibody/antigen-induced fluorescent signals were consecutively transferred from the biotin-streptavidin to CRISPR/Cas12a system. Third, the fluorescent signals were recorded by a smartphone-controlled handheld dark-box readout. Under optimization, detection limits in AFB1, BaP, and CAP were 0.00257, 4.971, and 794.6 fg/mL with wide linear ranges up to four orders of magnitude. Using urine, water, soybean oil, wheat, and peanuts as the complex matrix, we recorded high selectivity, considerable recovery, repeatability, and high consistency comparison to HPLC-MS/MS methods. This work promises a practical intelligent POCT platform for multiple targets in lipid-soluble and water-soluble matrices and could be extensively applied for healthcare, environment, and food safety.

Large-Scale Functionalized Metasurface-Based SARS-CoV-2 Detection and Quantification.

Plasmonic metasurfaces consist of metal-dielectric interfaces that are excitable at background and leakage resonant modes. The sharp and plasmonic excitation profile of metal-free electrons on metasurfaces at the nanoscale can be used for practical applications in diverse fields, including optoelectronics, energy harvesting, and biosensing. Currently, Fano resonant metasurface fabrication processes for biosensor applications are costly, need clean room access, and involve limited small-scale surface areas that are not easy for accurate sample placement. Here, we leverage the large-scale active area with uniform surface patterns present on optical disc-based metasurfaces as a cost-effective method to excite asymmetric plasmonic modes, enabling tunable optical Fano resonance interfacing with a microfluidic channel for multiple target detection in the visible wavelength range. We engineered plasmonic metasurfaces for biosensing through efficient layer-by-layer surface functionalization toward real-time measurement of target binding at the molecular scale. Further, we demonstrated the quantitative detection of antibodies, proteins, and the whole viral particles of SARS-CoV-2 with a high sensitivity and specificity, even distinguishing it from similar RNA viruses such as influenza and MERS. This cost-effective plasmonic metasurface platform offers a small-scale light-manipulation system, presenting considerable potential for fast, real-time detection of SARS-CoV-2 and pathogens in resource-limited settings.