LEPABE - Laboratory for Process Engineering, Environment, Biotechnology and Energy, Department of Chemical Engineering, Faculty of Engineering, University of Porto, Porto, Portugal.
CEFT - Transport Phenomena Research Center, Department of Chemical Engineering, Faculty of Engineering, University of Porto, Porto, Portugal.
Methods Mol Biol. 2021;2246:249-261. doi: 10.1007/978-1-0716-1115-9_16.
Suitable molecular methods for a faster microbial identification in food and clinical samples have been explored and optimized during the last decades. However, most molecular methods still rely on time-consuming enrichment steps prior to detection, so that the microbial load can be increased and reach the detection limit of the techniques.In this chapter, we describe an integrated methodology that combines a microfluidic (lab-on-a-chip) platform, designed to concentrate cell suspensions and speed up the identification process in Saccharomyces cerevisiae , and a peptide nucleic acid fluorescence in situ hybridization (PNA-FISH) protocol optimized and adapted to microfluidics. Microfluidic devices with different geometries were designed, based on computational fluid dynamics simulations, and subsequently fabricated in polydimethylsiloxane by soft lithography. The microfluidic designs and PNA-FISH procedure described here are easily adaptable for the detection of other microorganisms of similar size.
在过去几十年中，人们一直在探索和优化适用于食品和临床样本的更快微生物鉴定的分子方法。然而，大多数分子方法仍然依赖于在检测之前进行耗时的富集步骤，以便增加微生物负荷并达到技术的检测限。在本章中，我们描述了一种集成方法，该方法结合了微流控（芯片上实验室）平台，旨在浓缩细胞悬浮液并加速酿酒酵母的鉴定过程，以及优化和适应微流控的肽核酸荧光原位杂交（PNA-FISH）方案。基于计算流体动力学模拟设计了具有不同几何形状的微流控设备，然后通过软光刻在聚二甲基硅氧烷中制造。这里描述的微流控设计和 PNA-FISH 程序很容易适应检测其他类似大小的微生物。
