CRISPR-Cas12a-Mediated Hue-Recognition Lateral Flow Assay for Point-of-Need Detection of Salmonella.

Nucleic acid detection of pathogens in a point-of-need (PON) manner is of great significance yet remains challenging for sensitive and accurate visual discrimination. Here, we report a CRISPR-Cas12a-mediated lateral flow assay for PON detection of Salmonella typhimurium (S.ty) that is a prevailing pathogen disseminated through tainted food. The variation of the fluorescence color of the test line is exploited to interpret the results, enabling the discrimination between positive and negative samples on the basis of a hue-recognition mechanism. By leveraging the cleavage activity of Cas12a and hue-recognition readout, the assay facilitated by recombinase polymerase amplification can yield a visual detection limit of 1 copy µL-1 for S.ty genomic DNA within 1 h. The assay also displays a high specificity toward S.ty in fresh chicken samples, as well as a sensitivity 10-fold better than that of the commercial test strip. Moreover, a semiquantitative detection of S.ty ranging from 0 to 4 × 103 CFU/mL by the naked eye is made possible, thanks to the easily discernible color change of the test line. This approach provides an easy, rapid, accurate, and user-friendly solution for the PON detection of Salmonella and other pathogens.

Hue Recognition Competitive Fluorescent Lateral Flow Immunoassay for Aflatoxin M1 Detection with Improved Visual and Quantitative Performance.

Immunological detection of small molecules in a point-of-care (POC) format is of great significance yet remains challenging for accurate visual discrimination and quantitative analysis. Here, we report a novel hue recognition competitive fluorescent lateral flow immunoassay (HCLFIA) strip that allows both visual and quantitative detection of aflatoxin M1 (AFM1). The HCLFIA strip works on the basis of the ratiometric change of emission, arising from the overlap of fluorescence signals of two nanocomposites tagged with probe antibodies and coated antigens. A visually discernible orange-red-to-green fluorescence color change allows the naked eye semiquantitative readout of AFM1 around the threshold concentration (0.05 ng mL-1), yielding a visible detection limit of 0.02 ng mL-1. Moreover, using a custom smartphone-based device and color chart analysis, ultrasensitive quantitative detection of AFM1 can be achieved with a low limit of detection at 0.0012 ng mL-1, which is considerably better than those of the previously reported colorimetric and fluorescent strips. The accuracy performed in spiked milk samples ranged from 97.91 to 113.12% with a coefficient of variation below 7.8%, showing good consistency with the results from isotope dilution liquid chromatography-tandem mass spectrometry. Thanks to the unique hue recognition scheme, the HCLFIA strip holds great potential for POC detection of small molecules.

A solid-phase microextraction fiber coating based on magnetic covalent organic framework for highly efficient extraction of triclosan and methyltriclosan in environmental water and human urine samples.

Herein, we synthesized a kind of magnetic covalent organic framework nanohybrids (NiFe2O4@COF), and integrated it with polydimethyl siloxane and silicone rubber curing agent for solid phase microextraction (SPME) fiber coating. The fiber coating demonstrated a porous and uniform surface with the BET specific surface of 169.7 m2 g-1. As for seven environmental analytes, the NiFe2O4@COF-based SPME fiber coating gave the higher extraction recoveries for triclosan (TCS) and methyltriclosn (MTCS) than those of fenpropathrin, bifenthrin, permethrin, fenvalerate and deltamethrin. Several operational parameters were rigorously optimized, such as extraction temperature, extraction time, thermal desorption time, solution pH and salt effect. Combined with the GC-ECD detection, the newly developed microextraction method supplied the wide linear range of 0.1-1000 µg L-1 with the correlation coefficients of > 0.9995. The limits of detection (LODs) and limits of quantitation (LOQs) reached as low as 1-7 ng L-1 and 3.3-23 ng L-1, respectively. The intra-day and inter-day precisions in six replicates (n = 6 ) were < 3.55% and < 5.06%, respectively, and the fiber-to-fiber reproducibility (n = 3) was < 7.64%. To evaluate its feasibility in real samples, the fortified recoveries for TCS and MTCS, at low (0.2 µg L-1), middle (2.0 µg L-1) and high (20.0 µg L-1) levels, varied between 81.9% and 119.1% in tap, river and barreled waters as well as male, female and children urine samples. Especially, it is worth mentioning that the NiFe2O4@COF-based SPME coating fiber can be recycled for at least 150 times with nearly unchanged extraction efficiency. Moreover, the extraction recoveries by the as-fabricated fiber coating were much higher than those by three commercial fibers (PDMS, PDMS/DVB and PDMS/DVB/CAR). Overall, the NiFe2O4@COF-based SPME is a convenient, sensitive, efficient and "green" pretreatment method, thereby possessing important application prospects in trace monitoring of TCS-like pollutants in complex liquid matrices.