Institute of Bio- and Geosciences, IBG-1: Biotechnology, Forschungszentrum Jülich GmbH, Jülich, Germany.
Institute of Bio- and Geosciences, IBG-1: Biotechnology, Forschungszentrum Jülich GmbH, Jülich, Germany; Aachener Verfahrenstechnik (AVT), RWTH Aachen University, Aachen, Germany.
N Biotechnol. 2018 Dec 25;47:50-59. doi: 10.1016/j.nbt.2018.01.009. Epub 2018 Mar 14.
Microfluidics has enabled various research projects in the field of microbial single-cell analysis. In particular, single-use microfluidic cultivation devices combined with automated time-lapse imaging provide powerful approaches for analyzing microbial phenomena at the single-cell level. High spatiotemporal resolution facilitates individual cell identification and tracking, delivering detailed insights into areas like phenotypic population heterogeneity, which can be highly relevant to the fate of a microbial population and may negatively impact the efficiency of biotechnological fermentations. New tools need to be developed to access the origin of population heterogeneity and understand its functional role. In this study, we present a microfluidic device for batch cultivations inside picoliter-sized cultivation chambers that can be reversibly isolated from continuous medium supply. Therefore, the cultivation broth is simply replaced by a continuous flow of humidified air, removing any medium residue along the supply channels but preserving five picoliters of cultivation medium inside the cultivation chambers in a highly parallel manner. Living cells can grow inside our microfabricated batch chambers, which can accommodate up to several hundred cells. The chamber height approximately matches the diameter of a single cell, facilitating cell growth in monolayers that are ideal for image-based cell analysis. We successfully demonstrated the growth of Escherichia coli during continuous medium perfusion and batch cultivation conditions. As expected, the cells grew exponentially under continuous medium influx until the maximum chamber capacity was reached, but when they were cultivated under batch conditions, cellular growth underwent an exponential phase, followed by a stationary phase with obvious morphological changes.
微流控技术使得在微生物单细胞分析领域的各种研究项目成为可能。特别是，一次性微流控培养装置与自动化的时程成像相结合，为在单细胞水平上分析微生物现象提供了强大的方法。高时空分辨率有助于单个细胞的识别和跟踪，深入了解表型群体异质性等领域，这对于微生物群体的命运可能具有重要意义，并且可能会对生物技术发酵的效率产生负面影响。需要开发新的工具来了解群体异质性的起源及其功能作用。在这项研究中，我们提出了一种用于在皮升级别培养室内进行批量培养的微流控装置，该装置可与连续的培养基供应可逆隔离。因此，只需用加湿空气的连续流来替换培养肉汤，从而沿着供应通道去除任何培养基残留，但以高度并行的方式在培养室内保留 5 皮升的培养介质。活细胞可以在我们的微加工批量室内生长，每个室内可容纳多达几百个细胞。腔室的高度大致与单个细胞的直径相匹配，有利于细胞在单层中生长，这非常适合基于图像的细胞分析。我们成功地证明了大肠杆菌在连续介质灌注和批量培养条件下的生长。正如预期的那样，细胞在连续的介质流入下呈指数增长，直到达到最大腔室容量，但当它们在批量条件下培养时，细胞生长经历了指数期，然后进入具有明显形态变化的静止期。
