The double-membrane cell envelope of Gram-negative bacteria is a formidable barrier to intracellular antibiotic accumulation. A quantitative understanding of antibiotic transport in these cells is crucial for drug development, but this has proved elusive due to a dearth of suitable investigative techniques. Here we combine microfluidics and time-lapse auto-fluorescence microscopy to rapidly quantify antibiotic accumulation in hundreds of individual Escherichia coli cells. By serially manipulating the microfluidic environment, we demonstrated that stationary phase Escherichia coli, traditionally more refractory to antibiotics than growing cells, display reduced accumulation of the antibiotic ofloxacin compared to actively growing cells. Our novel microfluidic method facilitates the quantitative comparison of the role of the microenvironment versus that of the absence of key membrane transport pathways in cellular drug accumulation. Unlike traditional techniques, our assay is rapid, studying accumulation as the cells are dosed with the drug. This platform provides a powerful new tool for studying antibiotic accumulation in bacteria, which will be critical for the rational development of the next generation of antibiotics.

中文翻译：

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单细胞微流体有助于快速量化革兰氏阴性菌中的抗生素积累。

革兰氏阴性菌的双膜细胞包膜是细胞内抗生素积累的强大屏障。对这些细胞中抗生素转运的定量了解对于药物开发至关重要，但由于缺乏合适的研究技术，这已被证明是难以捉摸的。在这里，我们结合微流体和延时自荧光显微镜来快速量化数百个单独的大肠杆菌细胞中的抗生素积累。通过连续操纵微流体环境，我们证明了固定相大肠杆菌，传统上比生长的细胞对抗生素更难抵抗，与活跃生长的细胞相比，显示出抗生素氧氟沙星的积累减少。我们的新型微流体方法有助于定量比较微环境的作用与细胞药物积累中缺乏关键膜转运途径的作用。与传统技术不同的是，我们的检测速度很快，研究细胞在服用药物时的积累。该平台为研究细菌中抗生素积累提供了强大的新工具，这对于下一代抗生素的合理开发至关重要。