无需提取核酸的水凝胶环介导等温扩增技术用于超快速诊断粪便样本中的幽门螺旋杆菌

IF 5.7 2区化学 Q1 CHEMISTRY, ANALYTICAL

Analytica Chimica Acta Pub Date : 2024-11-07 DOI:10.1016/j.aca.2024.343384

Jinling Cui , Anning Tian , Haoran Wang , Yanan Yu , Jingwen Hao , Lei Wang , Chao Shi , Cuiping Ma

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引用次数: 0

摘要

背景幽门螺杆菌（Helicobacter pylori，H.pylori）感染引起的胃肠道疾病使幽门螺杆菌感染的准确检测变得更加重要。非侵入性方法，如分子诊断方法，已成为检测幽门螺杆菌的一种有前途的方法。粪便样本与环介导等温扩增（LAMP）相结合，显示出实时检测的潜在实用性。然而，在 LAMP 反应之前，需要复杂的核酸提取步骤来去除粪便样本中的大量扩增抑制剂。而且幽门螺杆菌的数量有限，导致检测反应时间长、灵敏度低。结果在本研究中，我们提出了一种无需提取的水凝胶 LAMP（hLAMP）超快速灵敏检测粪便样本中幽门螺杆菌的策略。水凝胶在简单的热裂解后与粪便样本结合，通过纳米定位在其纳米孔结构中形成扩增空间。纳米空间定位加速了 LAMP 反应。此外，由于水凝胶对复杂样品具有良好的抗抑制作用，这种基于 hLAMP 的方法可以在取样到检测结果的 40 分钟内特异、灵敏地检测出低至 100 CFU/mL 的幽门螺杆菌。整个过程包括 10 分钟的样品简单处理和 30 分钟的 LAMP 反应。意义这是首次应用水凝胶和 LAMP 系统结合检测粪便样本中的幽门螺杆菌。所开发的方法已在实际临床应用中得到验证，与常规组织病理学相比，准确率达到 88.9%。这也为幽门螺杆菌的诊断和预防提供了一个潜在的思路。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

Hydrogel loop-mediated isothermal amplification for ultra-fast diagnosis of Helicobacter pylori in stool samples without nucleic acid extraction

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Hydrogel loop-mediated isothermal amplification for ultra-fast diagnosis of Helicobacter pylori in stool samples without nucleic acid extraction

Background

The gastrointestinal diseases caused by Helicobacter pylori (H. pylori) infection made the accurate detection of H. pylori infection more important. Non-invasive methods, such as molecular diagnostic methods, had become a promising method for detection of H. pylori. Stool samples combined with loop-mediated isothermal amplification (LAMP), showed potential practicability for real-time detection. However, complex nucleic acid extraction steps were required to remove the large numbers of amplification inhibitors in stool samples before LAMP reaction. And the limited number of H. pylori made the detection with long reaction time and low sensitivity. The problems mentioned above were urgently to be solved.

Results

In this study, we proposed a strategy for ultra-rapid sensitive detection of H. pylori in stool samples by hydrogel LAMP (hLAMP) without extraction. The hydrogel was combined with stool samples after simple thermal cracking, and amplification spaces were formed in its nanopore structures by nano-localization. The LAMP reaction was accelerated by nano space-localization. Besides, this method based on hLAMP could specifically and sensitively detect as low as 100 CFU/mL H. pylori within 40 min from sampling to result due to good anti-inhibition effect on complex samples of hydrogel. The whole process involved sample simple disposal for 10 min and LAMP reaction for 30 min. Furthermore, the excellent anti-inhibition mechanism of hydrogel was discussed, and the mechanism of hydrogel accelerating LAMP was explored.

Significance

This is the first application of that hydrogel and LAMP systematically combined to detect H. pylori in stool samples. The developed method had been verified in actual clinical applications that the accuracy rate reached 88.9 % compared with routine histopathology. And it also provided a potential idea for the diagnosis and prevention of H. pylori.

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来源期刊

Analytica Chimica Acta 化学-分析化学

CiteScore

10.40

自引率

6.50%

发文量

1081

审稿时长

38 days

期刊介绍： Analytica Chimica Acta has an open access mirror journal Analytica Chimica Acta: X, sharing the same aims and scope, editorial team, submission system and rigorous peer review. Analytica Chimica Acta provides a forum for the rapid publication of original research, and critical, comprehensive reviews dealing with all aspects of fundamental and applied modern analytical chemistry. The journal welcomes the submission of research papers which report studies concerning the development of new and significant analytical methodologies. In determining the suitability of submitted articles for publication, particular scrutiny will be placed on the degree of novelty and impact of the research and the extent to which it adds to the existing body of knowledge in analytical chemistry.