Abstract
During the past 10 years, major developments in live-cell imaging methods have accompanied growing interest in the application of microfluidic techniques to biological imaging. The broad design possibilities of microfabrication and its relative ease of implementation have led to the development of a number of powerful imaging assays. Specifically, there has been great interest in the development of devices in which single cells can be followed in real-time over the course of several generations while the growth environment is changed. With standard perfusion chambers, the duration of a typical experiment is limited to one cell generation time. Using microfluidics, however, long-term imaging setups have been developed which can measure the effects of temporally controlled gene expression or pathway activation while tracking individual cells over the course of many generations. In this paper, we describe the details of fabricating such a microfluidic device for the purpose of long-term imaging of proliferating cells, the assembly of its individual components into a complete device, and then we give an example of how to use such a device to monitor real-time changes in gene expression in budding yeast. Our goal is to make this technique accessible to cell biology researchers without prior experience with microfluidic systems.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Chalfie M., Tu Y., Euskirchen G., Ward W.W. and Prasher D.C. (1994) Green fluorescent protein as a marker for gene expression. Science 263:802–5.
Tsien R.Y. (1998) The green fluorescent protein. Annu Rev Biochem 67:509–44.
Shaner N.C., Campbell R.E., Steinbach P.A., Giepmans B.N., Palmer A.E. and Tsien R.Y. (2004) Improved monomeric red, orange and yellow fluorescent proteins derived from Discosoma sp. red fluorescent protein. Nat Biotechnol 22:1567–72.
Goldman R.D. and Spector D.L. (2004) Live Cell Imaging. Cold Spring Harbour Laboratory Press, Woodbury, NY.
Voldman J., Gray M.L. and Schmidt M.A. (1999) Microfabrication in biology and medicine. Annu Rev Biomed Eng 1:401–25.
Whitesides G.M, Ostuni E., Takayama S., Jiang X. and Ingber D. (2001) Soft lithography in biology and biochemistry. Annu Rev Biomed Eng 3:335–73.
Sia S.K., Whitesides G.M. (2003) Microfluidic devices fabricated in poly(dimethylsiloxane) for biological studies. Electrophoresis 24(21): 3563–76.
Balaban N.Q., Merrin J., Chait R., Kowalik L. and Leibler S. (2004) Bacterial persistence as a phenotypic switch. Science 300(5690): 1622–5.
Paliwal S., Iglesias P.A., Campbell K., Hilioti Z., Groisman A. and Levchenko A. (2007) MAPK-mediated bimodal gene expression and adaptive gradient sensing in yeast. Nature 446(7131): 46–51.
Hersen P., McClean M.N., Mahadevan L., Ramanathan S. (2008) Signal processing by the HOG MAP kinase pathway. Proc Natl Acad Sci U S A 105(20):7165–70.
Bennett M.R., Pang W.L., Ostroff N.A., Baumgartner B.L., Nayak S., Tsimring L.S., Hasty J. (2008). Metabolic gene regulation in a dynamically changing environment. Nature 454(7208):1119–22.
McDonald J.C., Duffy D.C., Anderson J.R., Chiu J.T., Wu H., Schueller O.J.A., Whitesides G.M. (2000) Fabrication of microfluidic systems in poly(dimethylsiloxane). Electrophoresis 21:27–40.
Cookson S., Ostroff N., Pang W.L., Volfson D. and Hasty J. (2005) Monitoring dynamics of single-cell gene expression over multiple cell cycles. Mol Syst Biol 1:00024.
Lee P.J., Helman N.C., Lim W.A., and Hung P.J. (2008) A microfluidic system for dynamic yeast cell imaging. BioTechniques 44(1): 91–5
Charvin G., Cross F.R., Siggia E.D. (2008) A microfluidic device for temporally controlled gene expression and long-term fluorescent imaging in unperturbed dividing yeast cells. PLoS ONE 3:e1468.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2010 Humana Press, a part of Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Charvin, G., Oikonomou, C., Cross, F. (2010). Long-Term Imaging in Microfluidic Devices. In: Papkovsky, D. (eds) Live Cell Imaging. Methods in Molecular Biology, vol 591. Humana Press. https://doi.org/10.1007/978-1-60761-404-3_14
Download citation
DOI: https://doi.org/10.1007/978-1-60761-404-3_14
Published:
Publisher Name: Humana Press
Print ISBN: 978-1-60761-403-6
Online ISBN: 978-1-60761-404-3
eBook Packages: Springer Protocols