Abstract
Advancements in technology and computational power in recent years have directly impacted modern microscopy through improvements in light detection, imaging software platforms, as well as integration of complex hardware systems. These successes have allowed for mainstream utilization of previously complex microscopic techniques such as total internal reflection fluorescence (TIRF) microscopy, revealing many aspects of cell biology not previously appreciated. Through the restriction of illumination to areas of cell-coverslip interfaces in combination with modern detectors, TIRF microscopy allows researchers in the life sciences a glimpse of dynamic cellular phenomena with resolutions never before achieved.
This chapter provides a basic overview to the concept of TIRF microscopy and some considerations to setting up this technique in the lab.
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References
Alberts, D., and Papp, D. (Eds.) (1997) The Information Age: an Anthology on its Impact and Consequences. National Defense University Press, Washington, D.C.
Moore, G. (1965) Cramming more components onto integrated circuits. Electronics 38 114–17.
Liotta, L. A., and Kohn, E. C. (2001) The microenvironment of the tumour-host interface. Nature 411 375–9.
Aplin, A. E., Howe, A., Alahari, S. K., and Juliano, R. L. (1998) Signal transduction and signal modulation by cell adhesion receptors: the role of integrins, cadherins, immunoglobulin-cell adhesion molecules, and selectins. Pharmacol Rev 50 197–263.
Ward, Y., Wang, W., Woodhouse, E., Linnoila, I., Liotta, L., et al. (2001) Signal pathways which promote invasion and metastasis: critical and distinct contributions of extracellular signal-regulated kinase and Ral-specific guanine exchange factor pathways. Mol Cell Biol 21 5958–69.
Hanahan, D., and Weinberg, R. A. (2000) The hallmarks of cancer. Cell 100 57–70.
DeClerck, Y. A., Mercurio, A. M., Stack, M. S., Chapman, H. A., Zutter, M. M., et al. (2004) Proteases, extracellular matrix, and cancer: a workshop of the path B study section. Am J Pathol 164 1131–9.
Trache, A., and Meininger, G. A. (2008) Total internal reflection fluorescence (TIRF) microscopy. Curr Protoc Microbiol Chapter 2 Unit 2A 2 1-2A 2 22.
Kahraman, A., and Inal, M. E. (2002) Protective effects of quercetin on ultraviolet A light-induced oxidative stress in the blood of rat. J Appl Toxicol 22 303–9.
Stryer, L. (1995) DNA Structure, Replication, and Repair, in Biochemistry, W.H. Freeman and Company, New York, NY, pp. 811.
Lodish, H., Berk, A., Kaiser, C., Kreiger, M., Scott, M., et al. (2007) Molecular Cell Biology. W.H. Freeman and Company, New York, NY.
Serway, R., and Faughn, J. (1999) College Physics. Saunders College Publishing, Philadelphia, PA.
Ross, S., Schwartz, S., Fellers, T., and Davison, M. (2000) Total Internal Reflection Fluorescence (TIRF) Microscopy. September 27, 2009, http://www.microscopyu.com/articles/fluorescence/tirf/tirfintro.html
Byrne, G. D., Pitter, M. C., Zhang, J., Falcone, F. H., Stolnik, S., et al. (2008) Total internal reflection microscopy for live imaging of cellular uptake of sub-micron non-fluorescent particles. J Microsc 231 168–79.
Joselevitch, C., and Zenisek, D. (2009) Imaging exocytosis in retinal bipolar cells with TIRF microscopy. J Vis Exp 28, e1395, doi:10.3791/1305.
Young, I. T. (2001) Calibration: sampling density and spatial resolution. Curr Protoc Cytom Chapter 2 Unit 2 6.
Young, I. T. (1988) Sampling density and quantitative microscopy. Anal Quant Cytol Histol 10 269–75.
Blow, N. (2007) Cell migration: our protruding knowledge. Nature Methods 4 589–94.
Cohen, M., Kam, Z., Addadi, L., and Geiger, B. (2006) Dynamic study of the transition from hyaluronan- to integrin-mediated adhesion in chondrocytes. Embo J 25 302–11.
Huang, S., Lifshitz, L. M., Jones, C., Bellve, K. D., Standley, C., et al. (2007) Insulin Âstimulates membrane fusion and GLUT4 accumulation in clathrin coats on adipocyte plasma membranes. Mol Cell Biol 27 3456–69.
Huang, C., Rajfur, Z., Borchers, C., Schaller, M. D., and Jacobson, K. (2003) JNK phosphorylates paxillin and regulates cell migration. Nature 424 219–23.
Ewers, H., Smith, A. E., Sbalzarini, I. F., Lilie, H., Koumoutsakos, P., et al. (2005) Single-particle tracking of murine polyoma virus-like particles on live cells and artificial membranes. Proc Natl Acad Sci U S A 102 15110–5.
Hu, K., Ji, L., Applegate, K. T., Danuser, G., and Waterman-Storer, C. M. (2007) Differential transmission of actin motion within focal Âadhesions. Science 315 111–5.
Betzig, E., Patterson, G. H., Sougrat, R., Lindwasser, O. W., Olenych, S., et al. (2006) Imaging intracellular fluorescent proteins at nanometer resolution. Science 313 1642–5.
Rust, M. J., Bates, M., and Zhuang, X. (2006) Sub-diffraction-limit imaging by stochastic optical reconstruction microscopy (STORM). Nat Methods 3 793–5.
Eisenstein, M. (2006) Thinking big, seeing small. Nature 443 1019–20.
Shroff, H., Galbraith, C. G., Galbraith, J. A., and Betzig, E. (2008) Live-cell photoactivated localization microscopy of nanoscale adhesion dynamics. Nat Methods 5 417–23.
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Millis, B.A. (2012). Evanescent-Wave Field Imaging: An Introduction to Total Internal Reflection Fluorescence Microscopy. In: Espina, V., Liotta, L. (eds) Molecular Profiling. Methods in Molecular Biology, vol 823. Humana Press. https://doi.org/10.1007/978-1-60327-216-2_19
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DOI: https://doi.org/10.1007/978-1-60327-216-2_19
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