Ngamdee Tatchanun, Yin Lee Su, Vongpunsawad Sompong, Poovorawan Yong, Surareungchai Werasak, Lertanantawong Benchaporn
Department of Biotechnology, School of Bioresources and Technology, King Mongkut's University of Technology Thonburi, Bangkok, 10150, Thailand.
Faculty of Applied Sciences, AIMST University, Bedong, Kedah, 08100, Malaysia.
Anal Chim Acta. 2020 Oct 16;1134:10-17. doi: 10.1016/j.aca.2020.08.018. Epub 2020 Aug 21.
DNA strand displacement is an attractive, enzyme-free target hybridization strategy for nano-biosensing. The target DNA induces a strand displacement reaction by replacing the pre-hybridized strand that is labeled with gold nanoparticles (AuNPs). Thus, the amount of displaced-AuNP-labeled strand is proportional to the amount of target DNA in the sample. The use of a magnetogenosensing technique to isolate the target DNA allows for a simple, one-pot detection approach, which minimizes possible carry-over contamination and pipetting errors. We sought a proof-of-concept for this technology in its ability to detect DNA-equivalent of hepatitis E virus (HEV), which causes acute viral hepatitis for which rapid and simple diagnostic methods remain limited. Signal detection was done via visual observation, spectrophotometry, and electrochemistry. The sensor demonstrated good sensitivity with detection limits of 10 pM (visual), 10 pM (spectrophotometry) and 1 fM (electrochemical). This sensor also exhibited high specificity for real target amplicons and could discriminate between perfect and mismatched sequences. Lyophilized biosensor reagents stored at 4 °C, 25 °C, and outdoor ambient temperature, were stable for up to 90, 50, and 40 days, respectively. The integration of magnetic separation and target DNA-induced strand displacement reaction in a dry reagent form makes the sensing platform easy-to-use and suitable for field settings.
DNA链置换是一种用于纳米生物传感的有吸引力的、无酶的靶标杂交策略。靶标DNA通过取代预杂交的、标记有金纳米颗粒（AuNPs）的链来诱导链置换反应。因此，被置换的、标记有AuNP的链的量与样品中靶标DNA的量成正比。使用磁基因传感技术分离靶标DNA可实现一种简单的一锅法检测方法，该方法可将可能的残留污染和移液误差降至最低。我们寻求对该技术进行概念验证，以检测戊型肝炎病毒（HEV）的DNA等效物，戊型肝炎病毒会引发急性病毒性肝炎，而针对该疾病的快速简便诊断方法仍然有限。信号检测通过目视观察、分光光度法和电化学方法进行。该传感器表现出良好的灵敏度，检测限分别为10 pM（目视）、10 pM（分光光度法）和1 fM（电化学）。该传感器对实际靶标扩增子也表现出高特异性，并且能够区分完美序列和错配序列。冻干的生物传感器试剂分别在4℃、25℃和室外环境温度下储存，分别可稳定保存90天、50天和40天。以干燥试剂形式集成磁分离和靶标DNA诱导的链置换反应，使得传感平台易于使用且适用于现场环境。
