Our official English website, www.x-mol.net, welcomes your
feedback! (Note: you will need to create a separate account there.)
An Integrated Amplification-Free Digital CRISPR/Cas-Assisted Assay for Single Molecule Detection of RNA
ACS Nano ( IF 15.8 ) Pub Date : 2023-04-13 , DOI: 10.1021/acsnano.2c10143 Dou Wang 1 , Xuedong Wang 1 , Feidi Ye 2 , Jin Zou 2 , Jiuxin Qu 2 , Xingyu Jiang 1
ACS Nano ( IF 15.8 ) Pub Date : 2023-04-13 , DOI: 10.1021/acsnano.2c10143 Dou Wang 1 , Xuedong Wang 1 , Feidi Ye 2 , Jin Zou 2 , Jiuxin Qu 2 , Xingyu Jiang 1
Affiliation
|
Conventional nucleic acid detection technologies usually rely on amplification to improve sensitivity, which has drawbacks, such as amplification bias, complicated operation, high requirements for complex instruments, and aerosol pollution. To address these concerns, we developed an integrated assay for the enrichment and single molecule digital detection of nucleic acid based on a CRISPR/Cas13a and microwell array. In our design, magnetic beads capture and concentrate the target from a large volume of sample, which is 100 times larger than reported earlier. The target-induced CRISPR/Cas13a cutting reaction was then dispersed and limited to a million individual femtoliter-sized microwells, thereby enhancing the local signal intensity to achieve single-molecule detection. The limit of this assay for amplification-free detection of SARS-CoV-2 is 2 aM. The implementation of this study will establish a “sample-in-answer-out” single-RNA detection technology without amplification and improve the sensitivity and specificity while shortening the detection time. This research has broad prospects in clinical application.
中文翻译：
用于 RNA 单分子检测的集成无扩增数字 CRISPR/Cas 辅助测定
传统的核酸检测技术通常依靠扩增来提高灵敏度，存在扩增偏差、操作复杂、对仪器要求高、存在气溶胶污染等缺点。为了解决这些问题，我们开发了一种基于 CRISPR/Cas13a 和微孔阵列的核酸富集和单分子数字检测集成测定方法。在我们的设计中，磁珠从大量样品中捕获并浓缩目标，该样品比之前报道的样品大 100 倍。然后靶标诱导的CRISPR/Cas13a切割反应被分散并限制在100万个飞升大小的微孔中，从而增强局部信号强度以实现单分子检测。该检测方法对 SARS-CoV-2 的无扩增检测的极限是 2 aM。本研究的实施将建立一种“sample-in-answer-out”的无需扩增的单RNA检测技术，在缩短检测时间的同时提高灵敏度和特异性。该研究具有广阔的临床应用前景。
更新日期：2023-04-13
中文翻译：
用于 RNA 单分子检测的集成无扩增数字 CRISPR/Cas 辅助测定
传统的核酸检测技术通常依靠扩增来提高灵敏度，存在扩增偏差、操作复杂、对仪器要求高、存在气溶胶污染等缺点。为了解决这些问题，我们开发了一种基于 CRISPR/Cas13a 和微孔阵列的核酸富集和单分子数字检测集成测定方法。在我们的设计中，磁珠从大量样品中捕获并浓缩目标，该样品比之前报道的样品大 100 倍。然后靶标诱导的CRISPR/Cas13a切割反应被分散并限制在100万个飞升大小的微孔中，从而增强局部信号强度以实现单分子检测。该检测方法对 SARS-CoV-2 的无扩增检测的极限是 2 aM。本研究的实施将建立一种“sample-in-answer-out”的无需扩增的单RNA检测技术，在缩短检测时间的同时提高灵敏度和特异性。该研究具有广阔的临床应用前景。
京公网安备 11010802027423号