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Spinning and Fully Integrated Microdevice for Rapid Screening of Vancomycin-Resistant Enterococcus
ACS Sensors ( IF 8.2 ) Pub Date : 2021-07-22 , DOI: 10.1021/acssensors.1c00639
Thi Ngoc Diep Trinh 1 , Nae Yoon Lee 2
Affiliation  

This study introduces a spinning and fully integrated paper-based microdevice that can perform multiple functions, including DNA extraction, amplification, and colorimetric detection, for monitoring two major vancomycin-resistant Enterococci (VREs), which carry the vanA and vanB genes. The spinning microdevice is composed of a stationary part and a spinning part. The square-shaped stationary part has two zones: the lysis and reaction zones. The spinning part, which has a spin wheel-like shape, was inserted perpendicularly into the stationary part so that its two semicircles remained on the upper and lower parts. Sodium hydroxide-treated glass microfiber filter discs, inserted in the upper semicircle, were soaked in the lysis chambers by folding them toward the lysis zone to capture DNA in the lysis chambers. The captured DNA was transferred to the reaction chambers by folding the discs toward the reaction chambers. Water was added to the sodium hydroxide-treated glass microfiber filter discs to elute purified DNA into the reaction chambers. The upper semicircle was then unfolded, and the reaction chambers were sealed for subsequent loop-mediated isothermal amplification (LAMP) for 45 min. After the reaction, the spinning part was spun in the lysis zone direction to bring the lower semicircle, inserted with phenolphthalein-treated glass microfiber filter discs, toward the upper part of the stationary part. By folding it toward the reaction chambers, the lower semicircle came into contact with them and the phenolphthalein-treated glass microfiber filter discs were soaked in the reaction chambers and expressed color after 30 s. Based on the pH change during the LAMP reaction, the phenolphthalein-treated discs remained pink in the absence of target DNA, while those in contact with the positive samples turned colorless. A sensitive detection with a VRE limit of detection of 102 CFU/mL for tap water spiked with VRE carrying the vanA gene was achieved using this microdevice. Both VREs, carrying vanA and vanB genes, were successfully identified from tap water and contaminated equipment surfaces within 75 min. The introduced microdevice demonstrated a rapid, accurate, and sensitive performance for the environmental assessment of VRE contamination in resource-limited regions.

中文翻译:

用于快速筛选耐万古霉素肠球菌的旋转和全集成微装置

本研究介绍了一种旋转和完全集成的纸质微型设备,可以执行多种功能,包括 DNA 提取、扩增和比色检测,用于监测两种主要的耐万古霉素肠球菌(VRE),它们携带van A 和vanB基因。旋压微型器件由静止部分和旋压部分组成。方形固定部分有两个区域:裂解区和反应区。旋转部分呈旋转轮状,垂直插入静止部分,使其两个半圆保留在上部和下部。将经氢氧化钠处理的玻璃微纤维滤盘插入上半圆,通过将它们向裂解区折叠以捕获裂解室中的 DNA,从而将其浸泡在裂解室中。通过将圆盘向反应室折叠,将捕获的 DNA 转移到反应室。将水加入氢氧化钠处理过的玻璃微纤维滤盘中,以将纯化的 DNA 洗脱到反应室中。然后展开上半圆,将反应室密封,用于随后的环介导等温扩增 (LAMP) 45 分钟。反应结束后,纺丝部分沿裂解区方向旋转,使下半圆插入酚酞处理过的玻璃微纤维过滤盘,朝向静止部分的上部。通过将其向反应室折叠,下半圆与它们接触,酚酞处理过的玻璃微纤维滤片被浸泡在反应室中,并在 30 秒后呈现颜色。根据 LAMP 反应过程中的 pH 变化,酚酞处理的圆盘在没有目标 DNA 的情况下保持粉红色,而与阳性样品接触的圆盘则变为无色。VRE 检测限为 10 的灵敏检测 反应结束后,纺丝部分沿裂解区方向旋转,使下半圆插入酚酞处理过的玻璃微纤维过滤盘,朝向静止部分的上部。通过将其向反应室折叠,下半圆与它们接触,酚酞处理过的玻璃微纤维滤片被浸泡在反应室中,并在 30 秒后呈现颜色。根据 LAMP 反应过程中的 pH 变化,酚酞处理的圆盘在没有目标 DNA 的情况下保持粉红色,而与阳性样品接触的圆盘则变为无色。VRE 检测限为 10 的灵敏检测 反应结束后,纺丝部分沿裂解区方向旋转,使下半圆插入酚酞处理过的玻璃微纤维过滤盘,朝向静止部分的上部。通过将其向反应室折叠,下半圆与它们接触,酚酞处理过的玻璃微纤维滤片被浸泡在反应室中,并在 30 秒后呈现颜色。根据 LAMP 反应过程中的 pH 变化,酚酞处理的圆盘在没有目标 DNA 的情况下保持粉红色,而与阳性样品接触的圆盘则变为无色。VRE 检测限为 10 的灵敏检测 朝向静止部分的上部。通过将其向反应室折叠,下半圆与它们接触,酚酞处理过的玻璃微纤维滤片被浸泡在反应室中,并在 30 秒后呈现颜色。根据 LAMP 反应过程中的 pH 变化,酚酞处理的圆盘在没有目标 DNA 的情况下保持粉红色,而与阳性样品接触的圆盘则变为无色。VRE 检测限为 10 的灵敏检测 朝向静止部分的上部。通过将其向反应室折叠,下半圆与它们接触,酚酞处理过的玻璃微纤维滤片被浸泡在反应室中,并在 30 秒后呈现颜色。根据 LAMP 反应过程中的 pH 变化,酚酞处理的圆盘在没有目标 DNA 的情况下保持粉红色,而与阳性样品接触的圆盘则变为无色。VRE 检测限为 10 的灵敏检测 在没有目标 DNA 的情况下,酚酞处理的圆盘保持粉红色,而与阳性样品接触的圆盘则变为无色。VRE 检测限为 10 的灵敏检测 在没有目标 DNA 的情况下,酚酞处理的圆盘保持粉红色,而与阳性样品接触的圆盘则变为无色。VRE 检测限为 10 的灵敏检测使用该微型装置实现了2 CFU/mL 的自来水,其中掺入了带有van A 基因的VRE 。携带van A 和van B 基因的VRE在 75 分钟内从自来水和受污染的设备表面成功鉴定。引入的微型器件在资源有限地区的 VRE 污染环境评估中表现出快速、准确和灵敏的性能。
更新日期:2021-08-27
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