Zhao Liang, Zhao Zihao, Li Ning, Wang Xiayan
Center of Excellence for Environmental Safety and Biological Effects, Department of Chemistry, College of Chemistry and Life Science, Beijing University of Technology, Beijing, 100124, China.
Center of Excellence for Environmental Safety and Biological Effects, Department of Chemistry, College of Chemistry and Life Science, Beijing University of Technology, Beijing, 100124, China.
Talanta. 2025 May 1;286:127457. doi: 10.1016/j.talanta.2024.127457. Epub 2024 Dec 24.
Nucleic acid detection is considered the golden standard for diagnosing infectious diseases caused by various pathogens, including viruses, bacteria, and parasites. PCR and other amplification-based technologies are highly sensitive and specific, allowing for accurate detection and identification of low-level causative pathogens by targeting and amplifying their unique genetic segment (DNA or RNA). However, it is important to recognize that machinery-dependent diagnostic methods may only sometimes be available or practical in resource-limited settings, where direct implementation can be challenging. CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)-based diagnostics offer a promising alternative for nucleic acid detection. These methods provide gene sequence-specific targeting, multiplexing capability, rapid result disclosure, and ease of operation, making them suitable for point-of-care (POC) applications. CRISPR-Cas-based nucleic acid detection leverages the intrinsic gene-editing capabilities of CRISPR systems to detect specific DNA or RNA sequences with high precision, ensuring high specificity in identifying pathogens. When integrated with micro- and nano-technologies, CRISPR-based diagnostics gain additional benefits, including automated microfluidic processes, enhanced multiplexed detection, improved sensitivity through nanoparticle integration, and combined detection strategies. In this review, we analyze the motivations for tailoring the CRISPR-Cas system with microfluidic formats or nanoscale materials for nucleic acid biosensing and detection. We discuss and categorize current achievements in such systems, highlighting their differences, commonalities, and opportunities for addressing challenges, particularly for POC diagnostics. Micro- and nano-technologies can significantly enhance the practical utility of the CRISPR-Cas system, enabling more comprehensive diagnostic and surveillance capabilities. By integrating these technologies, CRISPR-based diagnostics can achieve higher levels of automation, sensitivity, and multiplexing, making them invaluable tools in the global effort to diagnose and control infectious diseases.
核酸检测被认为是诊断由各种病原体引起的传染病的金标准,这些病原体包括病毒、细菌和寄生虫。聚合酶链反应(PCR)和其他基于扩增的技术具有高度的敏感性和特异性,通过靶向和扩增病原体独特的基因片段(DNA或RNA),能够准确检测和鉴定低水平的致病病原体。然而,必须认识到,依赖仪器的诊断方法在资源有限的环境中可能并不总是可用或实用,在这些环境中直接实施可能具有挑战性。基于CRISPR(成簇规律间隔短回文重复序列)的诊断方法为核酸检测提供了一种有前景的替代方案。这些方法具有基因序列特异性靶向、多重检测能力、快速结果披露和操作简便等特点,使其适用于即时检测(POC)应用。基于CRISPR-Cas的核酸检测利用CRISPR系统固有的基因编辑能力来高精度检测特定的DNA或RNA序列,确保在识别病原体时具有高特异性。当与微纳技术集成时,基于CRISPR的诊断方法会获得更多优势,包括自动化微流控过程、增强的多重检测、通过纳米颗粒集成提高灵敏度以及联合检测策略。在本综述中,我们分析了将CRISPR-Cas系统与微流控形式或纳米材料相结合用于核酸生物传感和检测的动机。我们讨论并分类了此类系统的当前成果,突出它们的差异、共性以及应对挑战的机会,特别是对于即时检测诊断而言。微纳技术可以显著提高CRISPR-Cas系统的实际效用,实现更全面的诊断和监测能力。通过整合这些技术,基于CRISPR的诊断方法可以实现更高水平的自动化、灵敏度和多重检测,使其成为全球诊断和控制传染病努力中不可或缺的工具。