Department of Laboratory Medicine, The Affiliated Hospital of Southwest Medical University, Luzhou, 646000, PR China.
Department of Laboratory Medicine, The Affiliated Hospital of Southwest Medical University, Luzhou, 646000, PR China.
Talanta. 2022 Jun 1;243:123388. doi: 10.1016/j.talanta.2022.123388. Epub 2022 Mar 12.
Nucleic acid detection technology is now widely used in scientific research and clinical testing, such as infectious and genetic diseases screening, molecular diagnosis of tumors and pharmacogenomic research, which is also an important part of in vitro diagnostics (IVD). However, with the increasing requirements of diagnosis and treatment, existing nucleic acid detection technologies are facing challenges in dealing with the current problems (especially since the outbreak of coronavirus disease in 2019 (Covid-19)). Recently, Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) and CRISPR-associated protein (CRISPR/Cas)-based diagnostics have become a hot spot of attention. CRISPR/Cas has been developed as a molecular detection tool besides scientific research in biology and medicine fields, and some CRISPR-based products have already been translated. It is known as the "next-generation molecular diagnostic technology" because of its advantages such as easy design and accurate identification. CRISPR/Cas relies on pre-amplification of target sequences and subsequent detection of Cas proteins. Combining the CRISPR/Cas system with various isothermal nucleic acid amplification strategies can generate amplified detection signals, enrich low abundance molecular targets, improve the specificity and sensitivity of analysis, and develop point-of-care (POC) diagnostic techniques. In this review, we analyze the current status of CRISPR/Cas systems and isothermal amplification, report the advantages of combining the two and summarize the recent progress with the integration of both technologies with POC sensors in the nucleic acid field. In addition, the challenges and future prospects of CRISPR technology combined with isothermal amplification strategies in biosensing and clinical applications are discussed.
核酸检测技术目前广泛应用于科学研究和临床检测，例如传染病和遗传病筛查、肿瘤的分子诊断和药物基因组学研究，这也是体外诊断（IVD）的重要组成部分。然而，随着诊断和治疗要求的提高，现有的核酸检测技术在应对当前问题方面面临挑战（尤其是自 2019 年冠状病毒病（Covid-19）爆发以来）。最近，规律成簇间隔短回文重复序列（CRISPR）和 CRISPR 相关蛋白（CRISPR/Cas）为基础的诊断已成为关注的热点。CRISPR/Cas 除了在生物学和医学领域的科研之外，已经被开发成为一种分子检测工具，并且一些基于 CRISPR 的产品已经被转化。由于其易于设计和准确识别等优点，它被称为“下一代分子诊断技术”。CRISPR/Cas 依赖于目标序列的预扩增和随后的 Cas 蛋白检测。将 CRISPR/Cas 系统与各种等温核酸扩增策略相结合，可以产生扩增的检测信号，富集低丰度的分子靶标，提高分析的特异性和灵敏度，并开发即时检测（POC）诊断技术。在这篇综述中，我们分析了 CRISPR/Cas 系统和等温扩增的现状，报告了两者结合的优势，并总结了最近在核酸领域将两者技术与 POC 传感器集成的进展。此外，还讨论了 CRISPR 技术与等温扩增策略在生物传感和临床应用中的结合所面临的挑战和未来前景。
