An agar gel membrane-PDMS hybrid microfluidic device for long term single cell dynamic study

Significance of single cell measurements stems from the substantial temporal fluctuations and cell-cell variability possessed by individual cells. A major difficulty in monitoring surface non-adherent cells such as bacteria and yeast is that these cells tend to aggregate into clumps during growth, o...

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Bibliographic Details
Published in:Lab on a chip 2010-10, Vol.10 (20), p.2710-2719
Main Authors: Wong, Ieong, Atsumi, Shota, Huang, Wei-Chih, Wu, Tung-Yun, Hanai, Taizo, Lam, Miu-Ling, Tang, Ping, Yang, Jian, Liao, James C, Ho, Chih-Ming
Format: Article
Language:English
Subjects:
Agar
Agar - chemistry
Bacteria
Cell Culture Techniques - instrumentation
Devices
Dimethylpolysiloxanes - chemistry
Dynamics
Equipment Design
Equipment Failure Analysis
Flow Cytometry - instrumentation
Gels - chemistry
Membranes
Membranes, Artificial
Microfluidic Analytical Techniques - instrumentation
Microfluidics
Microorganisms
Nylons - chemistry
Tracking
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Summary:Significance of single cell measurements stems from the substantial temporal fluctuations and cell-cell variability possessed by individual cells. A major difficulty in monitoring surface non-adherent cells such as bacteria and yeast is that these cells tend to aggregate into clumps during growth, obstructing the tracking or identification of single-cells over long time periods. Here, we developed a microfluidic platform for long term single-cell tracking and cultivation with continuous media refreshing and dynamic chemical perturbation capability. The design highlights a simple device-assembly process between PDMS microchannel and agar membrane through conformal contact, and can be easily adapted by microbiologists for their routine laboratory use. The device confines cell growth in monolayer between an agar membrane and a glass surface. Efficient nutrient diffusion through the membrane and reliable temperature maintenance provide optimal growth condition for the cells, which exhibited fast exponential growth and constant distribution of cell sizes. More than 24 h of single-cell tracking was demonstrated on a transcription-metabolism integrated synthetic biological model, the gene-metabolic oscillator. Single cell morphology study under alcohol toxicity allowed us to discover and characterize cell filamentation exhibited by different E. coli isobutanol tolerant strains. We believe this novel device will bring new capabilities to quantitative microbiology, providing a versatile platform for single cell dynamic studies.
ISSN:1473-0197
1473-0189
DOI:10.1039/c004719h