This protocol describes the production and operation of a microfluidic dissection platform for long-term, high-resolution imaging of budding yeast cells. At the core of this platform is an array of micropads that trap yeast cells in a single focal plane. Newly formed daughter cells are subsequently washed away by a continuous flow of fresh culture medium. In a typical experiment, 50–100 cells can be tracked during their entire replicative lifespan. Apart from aging-related research, the microfluidic platform can also be a valuable tool for other studies requiring the monitoring of single cells over time. Here we provide step-by-step instructions on how to fabricate the silicon wafer mold, how to produce and operate the microfluidic device and how to analyze the obtained data. Production of the microfluidic dissection platform and setting up an aging experiment takes ∼7 h.
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We would like to thank M. Peter and U. Sauer for generously allowing us the use of the microscope; FIRST-CLA at ETH Zurich for the clean room facility during the development of this protocol; L. Lee for initial discussions; and N. Thayer, A. Papagiannakis and A. Meinema for helpful comments on the manuscript. Funding from the NWO-funded Groningen Systems Biology Center for Energy Metabolism and Ageing is acknowledged.
The authors declare no competing financial interests.
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Huberts, D., Sik Lee, S., González, J. et al. Construction and use of a microfluidic dissection platform for long-term imaging of cellular processes in budding yeast. Nat Protoc 8, 1019–1027 (2013). https://doi.org/10.1038/nprot.2013.060
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