Key Laboratory of East China Sea Fishery Resources Exploitation, Ministry of Agriculture, East China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Shanghai, 200090, China.
Laboratory of Physical Chemistry, Université Felix Houphouet-Boigny, 22 BP 582, Abidjan 22, Côte d'Ivoire.
Mikrochim Acta. 2017 Dec 7;185(1):35. doi: 10.1007/s00604-017-2594-4.
Loop-mediated isothermal amplification (LAMP) eradicates the need of thermocycler in DNA amplification. Signals are usually obtained via fluorometry or turbidimetry, but such methods need improvement in order to become more effortless and reliable. The authors describe a set of six specific primers targeting the species-specific tlh gene of Vibrio parahaemolyticus which were used in accelerated LAMP reaction. Gold nanoparticles (AuNPs) were functionalized with streptavidin (Avidin-AuNPs), and engineered to signal the LAMP reaction. Two of the loop primers for LAMP were biotinylated and then can produce a DNA that can cause clusterization of Avidin-AuNPs based on the formation of avidin-biotin complex. This leads to a color change of the solution from red to blue. Amplification is completed within 30 min and can be visually detected within 5 min. The detection limit of the method is found to be 8.6 cfu per reaction. This visual detection scheme does not require any fluorescent reagents and detection instruments. Conceivably, the method has a wide scope because such Avidin-AuNPs can be used as nanoprobes for a variety of other LAMP products. This rapid and universal strategy holds promise in point of care testing and food testing, particularly in resource-limited regions. Graphical abstract Six specific primers (two of them are biotinylated) were used to realize the accelerated Loop-Mediated Isothermal Amplification. Streptavidin modified gold nanoparticles (Avidin-AuNPs) cluster on the DNA products, leading to the apparent change of color from red to blue, which is readily identified even by unaided eye.
环介导等温扩增（LAMP）消除了 DNA 扩增对热循环仪的需求。信号通常通过荧光计或浊度计获得，但这些方法需要改进，以变得更加轻松和可靠。作者描述了一组针对副溶血性弧菌种特异性 tlh 基因的六个特异性引物，用于加速 LAMP 反应。金纳米粒子（AuNPs）被链霉亲和素（avidin-AuNPs）功能化，并被设计用于信号 LAMP 反应。LAMP 的两个环引物被生物素化，然后可以产生一种 DNA，基于亲和素-生物素复合物的形成，导致avidin-AuNPs 的聚集。这导致溶液的颜色从红色变为蓝色。扩增在 30 分钟内完成，可在 5 分钟内进行目视检测。该方法的检测限发现为每个反应 8.6cfu。这种目视检测方案不需要任何荧光试剂和检测仪器。可以想象，该方法具有广泛的应用范围，因为这种avidin-AuNPs 可以用作各种其他 LAMP 产品的纳米探针。这种快速和通用的策略有望在即时检测和食品检测中得到应用，特别是在资源有限的地区。示意图六组特异性引物（其中两组被生物素化）用于实现加速的环介导等温扩增。链霉亲和素修饰的金纳米粒子（avidin-AuNPs）在 DNA 产物上聚集，导致颜色从红色明显变为蓝色，即使没有肉眼也很容易识别。
