Zheng Laibao, Zheng Chaochuan, Wang Weiwei, Huang Fuyuan, Jiang Yelin, Lu Jiahai, Lou Yongliang
Wenzhou Key Laboratory of Sanitary Microbiology, Key Laboratory of Laboratory Medicine, Ministry of Education, School of Laboratory Medicine and Life Sciences, Wenzhou Medical University, Wenzhou 325035, China.
National Clinical Research Center for Ocular Diseases, Eye Hospital, Wenzhou Medical University, Wenzhou 325027, China.
Colloids Surf B Biointerfaces. 2025 Jun;250:114541. doi: 10.1016/j.colsurfb.2025.114541. Epub 2025 Jan 30.
Pathogenic bacteria, such as Pseudomonas aeruginosa, pose significant threats to public health due to their multidrug resistance and association with severe infections. Rapid and reliable detection methods are crucial for timely treatment and effective infection control, especially in resource-limited settings. In this study, we developed a CRISPR/Cas12a-based colorimetric biosensor that leverages Cas12a's trans-cleavage activity to release left single-stranded DNA (lDNA). The released lDNA facilitates hybridization with clDNA-functionalized gold nanoparticles (AuNPs), resulting in a visible color change. The biosensor achieved a detection limit of 10 CFU/reaction for P. aeruginosa within 2 hours, with excellent specificity and robustness, as validated in spiked sputum and blood samples. Clinical testing using 32 blood samples (13 positive, 19 negative) confirmed its high diagnostic accuracy, achieving an AUC of 1 in ROC curve analysis. The platform's simplicity, robustness, and programmability suggest its broad potential for rapid infectious disease diagnostics, particularly in low-resource settings.
致病性细菌，如铜绿假单胞菌，由于其多重耐药性以及与严重感染的关联，对公共卫生构成了重大威胁。快速且可靠的检测方法对于及时治疗和有效的感染控制至关重要，尤其是在资源有限的环境中。在本研究中，我们开发了一种基于CRISPR/Cas12a的比色生物传感器，该传感器利用Cas12a的反式切割活性来释放左侧单链DNA（lDNA）。释放出的lDNA有助于与经clDNA功能化的金纳米颗粒（AuNP）杂交，从而导致可见的颜色变化。该生物传感器在2小时内对铜绿假单胞菌的检测限达到10 CFU/反应，具有出色的特异性和稳健性，在加标的痰液和血液样本中得到了验证。使用32份血液样本（13份阳性，19份阴性）进行的临床试验证实了其高诊断准确性，在ROC曲线分析中AUC为1。该平台的简单性、稳健性和可编程性表明其在快速传染病诊断方面具有广阔的潜力，特别是在资源匮乏的环境中。
