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- Title
Cascaded signal amplification strategy for ultra-specific, ultra-sensitive, and visual detection of Shigella flexneri.
- Authors
Shi, Yaoqiang; Tan, Qi; Gong, Tao; Li, Qing-yuan; Zhu, Ya; Duan, Xiaoqiong; Yang, Chunhui; Ding, Jia-wei; Li, Shilin; Xie, He; Li, Yujia; Chen, Limin
- Abstract
Pathogen infections including Shigella flexneri have posed a significant threat to human health for numerous years. Although culturing and qPCR were the gold standards for pathogen detection, time-consuming and instrument-dependent restrict their application in rapid diagnosis and economically less-developed regions. Thus, it is urgently needed to develop rapid, simple, sensitive, accurate, and low-cost detection methods for pathogen detection. In this study, an immunomagnetic beads-recombinase polymerase amplification-CRISPR/Cas12a (IMB-RPA-CRISPR/Cas12a) method was built based on a cascaded signal amplification strategy for ultra-specific, ultra-sensitive, and visual detection of S. flexneri in the laboratory. Firstly, S. flexneri was specifically captured and enriched by IMB (Shigella antibody-coated magnetic beads), and the genomic DNA was released and used as the template in the RPA reaction. Then, the RPA products were mixed with the pre-loaded CRISPR/Cas12a for fluorescence visualization. The results were observed by naked eyes under LED blue light, with a sensitivity of 5 CFU/mL in a time of 70 min. With no specialized equipment or complicated technical requirements, the IMB-RPA-CRISPR/Cas12a diagnostic method can be used for visual, rapid, and simple detection of S. flexneri and can be easily adapted to monitoring other pathogens.
- Subjects
SHIGELLA flexneri; SHIGELLOSIS; BLUE light; CRISPRS; POLYMERASES; PRODUCT mixes
- Publication
Microchimica Acta, 2024, Vol 191, Issue 5, p1
- ISSN
0026-3672
- Publication type
Article
- DOI
10.1007/s00604-024-06309-0