This title appears in the Scientific Report :
2013
Please use the identifier:
http://hdl.handle.net/2128/5678 in citations.
Please use the identifier: http://dx.doi.org/10.3791/50560 in citations.
Microfluidic Picoliter Bioreactor for Microbial Single-cell Analysis: Fabrication, System Setup, and Operation
Microfluidic Picoliter Bioreactor for Microbial Single-cell Analysis: Fabrication, System Setup, and Operation
In this protocol the fabrication, experimental setup and basic operation of the recently introduced microfluidic picoliter bioreactor (PLBR) is described in detail. The PLBR can be utilized for the analysis of single bacteria and microcolonies to investigate biotechnological and microbiological rela...
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Personal Name(s): | Grünberger, Alexander (Corresponding author) |
---|---|
Probst, Christopher / Heyer, Antonia / Wiechert, Wolfgang / Frunzke, Julia / Kohlheyer, Dietrich | |
Contributing Institute: |
Biotechnologie; IBG-1 |
Published in: | Journal of visualized experiments, 82 (2013) S. 50560 |
Imprint: |
[S.l.]
JoVE
2013
|
DOI: |
10.3791/50560 |
PubMed ID: |
24336165 |
Document Type: |
Journal Article |
Research Program: |
ohne Topic |
Link: |
OpenAccess |
Publikationsportal JuSER |
Please use the identifier: http://dx.doi.org/10.3791/50560 in citations.
In this protocol the fabrication, experimental setup and basic operation of the recently introduced microfluidic picoliter bioreactor (PLBR) is described in detail. The PLBR can be utilized for the analysis of single bacteria and microcolonies to investigate biotechnological and microbiological related questions concerning, e.g. cell growth, morphology, stress response, and metabolite or protein production on single-cell level. The device features continuous media flow enabling constant environmental conditions for perturbation studies, but in addition allows fast medium changes as well as oscillating conditions to mimic any desired environmental situation. To fabricate the single use devices, a silicon wafer containing sub micrometer sized SU-8 structures served as the replication mold for rapid polydimethylsiloxane casting. Chips were cut, assembled, connected, and set up onto a high resolution and fully automated microscope suited for time-lapse imaging, a powerful tool for spatio-temporal cell analysis. Here, the biotechnological platform organism Corynebacterium glutamicum was seeded into the PLBR and cell growth and intracellular fluorescence were followed over several hours unraveling time dependent population heterogeneity on single-cell level, not possible with conventional analysis methods such as flow cytometry. Besides insights into device fabrication, furthermore, the preparation of the preculture, loading, trapping of bacteria, and the PLBR cultivation of single cells and colonies is demonstrated. These devices will add a new dimension in microbiological research to analyze time dependent phenomena of single bacteria under tight environmental control. Due to the simple and relatively short fabrication process the technology can be easily adapted at any microfluidics lab and simply tailored towards specific needs. |