A Highly Sensitive CRISPR‐Empowered Surface Plasmon Resonance Sensor for Diagnosis of Inherited Diseases with Femtomolar‐Level Real‐Time Quantification

作者：Fei Zheng, Zhi Chen, Jingfeng Li, Rui Wu, Bin Zhang, Guohui Nie, Zhongjian Xie, Han Zhang · 发表于：Advanced Science · 年份：2022 · DOI：10.1002/advs.202105231 · 被引用次数：296 · 研究领域：CRISPR and Genetic Engineering、Advanced biosensing and bioanalysis techniques、Biosensors and Analytical Detection

The clustered regularly interspaced short palindromic repeats (CRISPR) molecular system has emerged as a promising technology for the detection of nucleic acids. Herein, the development of a surface plasmon resonance (SPR) sensor that is functionalized with a layer of locally grown graphdiyne film, achieving excellent sensing performance when coupled with catalytically deactivated CRISPR-associated protein 9 (dCas9), is reported. dCas9 protein is immobilized on the sensor surface and complexed with a specific single-guide RNA, enabling the amplification-free detection of target sequences within genomic DNA. The sensor, termed CRISPR-SPR-Chip, is used to successfully analyze recombinant plasmids with only three-base mutations with a limit of detection as low as 1.3 fM. Real-time monitoring CRISPR-SPR-Chip is used to analyze clinical samples of patients with Duchenne muscular dystrophy with two exon deletions, which are detected without any pre-amplification step, yielding significantly positive results within 5 min. The ability of this novel CRISPR-empowered SPR (CRISPR-eSPR) sensing platform to rapidly, precisely, sensitively, and specifically detect a target gene sequence provides a new on-chip optic approach for clinical gene analysis.