Sutlief Arin L, Valquier-Flynn Helena, Wilson Christina, Perez Marco, Kleinschmidt Hunter, Schofield Brett J, Delmain Elizabeth, Holmes Andrea E, Wentworth Christopher D
Department of Chemistry, Doane University;
Department of Chemistry, Doane University.
J Vis Exp. 2019 Jan 16(143). doi: 10.3791/58926.
A higher-throughput microfluidic in vitro bioreactor coupled with fluorescence microscopy has been used to study bacterial biofilm growth and morphology, including Pseudomonas aeruginosa (P. aeruginosa). Here, we will describe how the system can be used to study the growth kinetics and the morphological properties such as the surface roughness and textural entropy of P. aeruginosa strain PA01 that expresses an enhanced green fluorescent protein (PA01-EGFP). A detailed protocol will describe how to grow and seed PA01-EGFP cultures, how to set up the microscope and autorun, and conduct the image analysis to determine growth rate and morphological properties using a variety of shear forces that are controlled by the microfluidic device. This article will provide a detailed description of a technique to improve the study of PA01-EGFP biofilms which eventually can be applied towards other strains of bacteria, fungi, or algae biofilms using the microfluidic platform.
一种高通量微流控体外生物反应器与荧光显微镜联用，已被用于研究细菌生物膜的生长和形态，包括铜绿假单胞菌（P. aeruginosa）。在此，我们将描述该系统如何用于研究表达增强型绿色荧光蛋白的铜绿假单胞菌PA01菌株（PA01-EGFP）的生长动力学以及诸如表面粗糙度和纹理熵等形态学特性。一份详细的方案将描述如何培养和接种PA01-EGFP培养物，如何设置显微镜并自动运行，以及如何进行图像分析以确定生长速率和形态学特性，该分析使用由微流控装置控制的各种剪切力。本文将详细描述一种改进PA01-EGFP生物膜研究的技术，该技术最终可应用于使用微流控平台研究其他细菌、真菌或藻类生物膜菌株。
