Experimental Biophysics & Applied Nanoscience, Bielefeld University, Universitaetsstr. 25, 33615 Bielefeld, Germany.
J Biotechnol. 2010 Sep 15;149(4):280-8. doi: 10.1016/j.jbiotec.2010.06.003. Epub 2010 Jun 25.
Life cell imaging of bacterial cells over long times is very challenging because of the small dimensions and the need for a liquid environment assuring cell viability. In order to obtain space- and time-resolved information about protein dynamics, high resolution time-lapse fluorescence images (TLFI) of single bacterial cells were recorded in a poly(dimethylsiloxane) (PDMS) microfluidic chip. A new gradient coating technique was applied to ensure cell loading. As a proof-of-concept, we monitored the evenly distributed cytoplasmic protein GcrA as well as the asymmetric localization of the DivK protein in cells of S. meliloti over at least two division cycles. Localization of DivK was characterized by dividing each bacterial cell into 4 sections with dimensions closely above the optical limit of resolution. This approach of generating spatio-temporal resolved information of protein dynamics in single bacterial cells is applicable to many problems.
对细菌细胞进行长时间的生命细胞成像非常具有挑战性，因为细菌细胞体积小，并且需要确保细胞活力的液体环境。为了获取关于蛋白质动力学的时空分辨信息，在聚二甲基硅氧烷（PDMS）微流控芯片中记录了单个细菌细胞的高分辨率延时荧光图像（TLFI）。应用了一种新的梯度涂层技术来确保细胞加载。作为概念验证，我们监测了均匀分布的细胞质蛋白 GcrA 以及 S. meliloti 细胞中 DivK 蛋白的不对称定位，至少两个分裂周期。DivK 的定位通过将每个细菌细胞划分为 4 个部分来实现，其尺寸接近光学分辨率的极限。这种在单个细菌细胞中生成蛋白质动力学时空分辨信息的方法适用于许多问题。
