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We combined a CRISPR/Cas12a system with a hybridization chain reaction (HCR) to develop an ultrasensitive magnetic relaxation switching (MRS) biosensor for detecting viable Salmonella typhimurium ( S. typhimurium ). Magnetic nanoparticles of two sizes (30 and 1000 nm: MNP 30 and MNP 1000 , respectively) were coupled through HCR. The S. typhimurium gene-activated CRISPR/Cas12a system released MNP 30 from the MNP 1000 -HCR-MNP 30 complex through a trans-cleavage reaction. After magnetic separation, released MNP 30 was collected from the supernatant and served as a transverse relaxation time (T 2 ) signal probe. Quantitative detection of S. typhimurium is achieved by establishing a linear relationship between the change in T 2 and the target gene. The biosensor's limit of detection was 77 CFU/mL (LOD = 3 S / M , S = 22.30, M = 0.87), and the linear range was 10 2 -10 8 CFU/mL. The accuracy for detecting S. typhimurium in real samples is comparable to that of qPCR. Thus, this is a promising method for the rapid and effective detection of foodborne pathogens.

Deoxynivalenol is one of the most widely distributed mycotoxins in cereals and poses tremendous threats to the agricultural environment and public health. Therefore, it is particularly important to develop sensitive and interference-resistant deoxynivalenol analysis methods. Here, we establish a "Lollipop" particle counting immunoassay (LPCI) based on antigen-powered CRISPR-Cas12a dual signal amplification. LPCI achieves high sensitivity and accuracy through antigen-powered CRISPR-Cas dual signal amplification combined with particle counting immunoassay. This strategy not only broadens the applicability of the CRISPR-Cas system in the field of non-nucleic acid target detection; it also improves the sensitivity of particle counting immunoassay. The introduction of a polystyrene "lollipop" immunoassay carrier further enables efficiently simultaneous pre-treatment of multiple samples and overcomes complex matrix interference in real samples. The linear detection range of LPCI for deoxynivalenol was 0.1-500 ng/mL with a detection limit of 0.061 ng/mL. The platform greatly broadens the scope of the CRISPR-Cas sensor for the detection of non-nucleic acid hazards in the environment and food samples., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Deoxynivalenol (DON) is highly toxic to the environment and human health. It is important to detect DON with ultra-high sensitivity, ease of operation, and low cost. Inspired by the excellent stability and biocompatibility of polydopamine, a universal, portable and ultra-sensitive pipet immunoassay platform was reported for DON detection based on dopamine self-polymerization (polydopamine coating and polydopamine nanoparticles). The polydopamine coating acted as an effective strategy for biomolecule immobilization on the pipet to improve the coating efficiency that significantly reduced the required concentration of biomolecules. Performing the ELISA in pipets saved nearly 67% of the antigen amount and 83% of the antibody amount, which reduced the detection cost and simplified the experimental steps. The dual signal amplification in this pipet immunoassay enabled ultra-high sensitivity. Polydopamine nanoparticles acted as the enrichment carrier of horseradish peroxidase-goat anti-mouse IgG for the first-round signal amplification, followed by the tyramine-mediated loading of streptavidin-HRP for the second-round signal amplification. The dual-enriched HRP catalyzed the color-developing substrate to achieve highly sensitive colorimetric DON detection with a limit of detection of 0.435 ng/mL., (Copyright © 2022 Elsevier B.V. All rights reserved.)

MicroRNAs (miRNAs) constitute novel biomarkers for various diseases. Accurate and quantitative analysis of miRNA expression is critical for biomedical research and clinical theranostics. In this study, a method was developed for sensitive and specific detection of miRNAs via dual signal amplification based on duplex specific nuclease (DSN) and hybridization chain reaction (HCR). A reporter probe (RP), comprising recognition sequence (3' end modified with biotin) for a target miRNA of miR-21 and capture sequence (5' end modified with Fam) for HCR product, was designed and synthesized. HCR was initiated by partial sequence of initiator probe (IP), the other part of which can hybridize with capture sequence of RP, and was assembled by hairpin probes modified with biotin (H1-bio and H2-bio). A miR-21 triggered cyclical DSN cleavage of RP, which was immobilized to a streptavidin (SA) coated magnetic bead (MB). The released Fam labeled capture sequence then hybridized with the HCR product to generate a detectable dsDNA. This polymer was then dropped on lateral flow strip and positive result was observed. The proposed method allowed quantitative sequence-specific detection of miR-21 (with a detection limit of 2.1 fM, S/N = 3) in a dynamic range from 100 fM to 100 pM, with an excellent ability to discriminate differences in miRNAs. The method showed acceptable testing recoveries for the determination of miRNAs in serum.