The development of a biosensing platform with high sensitivity, high specificity, and low cost for the detection of biomarkers, especially one that is programmable and universal, is critical for disease surveillance and diagnosis, yet it remains a difficulty. Herein, we combined the clustered regularly interspaced short palindromic repeats (CRISPR)/Cas system with a fluorescent label-free biosensor platform for sensitive and specific detection of disease-related protein, small molecule and nucleic acid. In this strategy, we designed an exonuclease III-mediated target cycle and released a universal trigger chain to stimulate the enzyme activity of CRISPR/Cas12a for additional signal amplification. The hydrolysis of ssDNA-templated silver nanoclusters (ssDNA-Ag NCs) as the reporter probe resulted in a significant decrease of fluorescence intensity. This biosensing platform can be flexibly used to the sensitive and specific determination of protein, small molecule, or microRNA in biological samples by simply transforming the target recognized sequences in the DNA hairpin. In this work, a new label-free sensing system used the fluorescent ssDNA-Ag NCs as the signal output does not need to be marked in advance and has no background signal. In addition, the method has the advantages of low cost, simple operation and high speed, and provides an innovative idea for the development of a powerful clinical diagnosis tool.

开发一种用于检测生物标志物的高灵敏度、高特异性和低成本的生物传感平台，尤其是可编程和通用的生物传感平台，对于疾病监测和诊断至关重要，但仍然是一个难题。在此，我们将成簇的规则间隔短回文重复序列 (CRISPR)/Cas 系统与无荧光标记的生物传感器平台相结合，用于对疾病相关的蛋白质、小分子和核酸进行灵敏和特异性检测。在该策略中，我们设计了一个外切核酸酶 III 介导的靶循环，并释放了一条通用触发链来刺激 CRISPR/Cas12a 的酶活性，从而实现额外的信号放大。作为报告探针的 ssDNA 模板化银纳米簇 (ssDNA-Ag NCs) 的水解导致荧光强度显着降低。该生物传感平台可灵活用于生物样品中蛋白质、小分子或 microRNA 的灵敏和特异性测定，只需简单地转换 DNA 发夹中的目标识别序列即可。在这项工作中，一种新的无标记传感系统使用荧光 ssDNA-Ag NCs 作为信号输出，不需要预先标记，也没有背景信号。此外，该方法成本低、操作简单、速度快，为开发强大的临床诊断工具提供了创新思路。一种新的无标记传感系统使用荧光 ssDNA-Ag NCs 作为信号输出不需要预先标记并且没有背景信号。此外，该方法成本低、操作简单、速度快，为开发强大的临床诊断工具提供了创新思路。一种新的无标记传感系统使用荧光 ssDNA-Ag NCs 作为信号输出不需要预先标记并且没有背景信号。此外，该方法成本低、操作简单、速度快，为开发强大的临床诊断工具提供了创新思路。