Fletcher Matthew P, Diggle Stephen P, Cámara Miguel, Williams Paul
Centre for Biomolecular Sciences, School of Life Sciences, University of Nottingham, Nottingham, NG7 2RD, UK.
Methods Mol Biol. 2018;1673:25-34. doi: 10.1007/978-1-4939-7309-5_2.
2-Alkyl-4-quinolones (AQs) such as 2-heptyl-3-hydroxy-4-quinolone (PQS) and 2-heptyl-4-hydroxyquinoline (HHQ) are quorum-sensing signal molecules. Here we describe two methods for AQ detection and quantification that employ thin-layer chromatography (TLC) and microtiter plate assays in combination with a lux-based Pseudomonas aeruginosa AQ biosensor strain. For TLC detection, organic solvent extracts of bacterial cells or spent culture supernatants are chromatographed on TLC plates, which are then dried and overlaid with the AQ biosensor. After detection by the bioreporter, AQs appear as both luminescent and green (from pyocyanin) spots. For the microtiter assay, either spent bacterial culture supernatants or extracts are added to a growth medium containing the AQ biosensor. Light output by the bioreporter correlates with the AQ content of the sample. The assays described are simple to perform, do not require sophisticated instrumentation, and are highly amenable to screening large numbers of bacterial samples.
2-烷基-4-喹诺酮（AQs），如2-庚基-3-羟基-4-喹诺酮（PQS）和2-庚基-4-羟基喹啉（HHQ），是群体感应信号分子。在此，我们描述了两种用于AQ检测和定量的方法，这些方法结合了薄层色谱（TLC）和微量滴定板分析，并使用基于lux的铜绿假单胞菌AQ生物传感器菌株。对于TLC检测，将细菌细胞或用过的培养上清液的有机溶剂提取物在TLC板上进行色谱分析，然后干燥并覆盖上AQ生物传感器。经生物报告器检测后，AQs呈现为发光和绿色（来自绿脓菌素）斑点。对于微量滴定分析，将用过的细菌培养上清液或提取物添加到含有AQ生物传感器的生长培养基中。生物报告器发出的光输出与样品中的AQ含量相关。所描述的分析方法操作简单，不需要复杂的仪器，并且非常适合筛选大量细菌样品。
