Abstract
In recent years three powerful optical imaging techniques have emerged that provide nanometer-scale information about the topography of membrane surfaces, whether cellular or artificial: intermembrane fluorescence resonance energy transfer (FRET), fluorescence interference contrast microscopy (FLIC), and reflection interference contrast microscopy (RICM). In intermembrane FRET, the sharp distance dependence of resonant energy transfer between fluorophores allows topographic measurements in the Ångstrom to few-nanometer range. In FLIC and RICM, interference between light from a membrane (either from fluorescent probes, or reflected illumination) and light reflected by a planar substrate provide spatial sensitivity in the few to hundreds of nanometer range, with few-nanometer resolution. All of these techniques are fairly easy to implement. We discuss the physics and optics behind each of these tools, as well as practical concerns regarding their uses. We also provide examples of their application in imaging molecular-scale structures at intermembrane junctions.
Similar content being viewed by others
References
Simon, S. I. and Goldsmith, H. L. (2002) Leukocyte adhesion dynamics in shear flow. Ann. Biomed. Eng. 30, 315–332.
Dworak, H. A. and Sink, H. (2002) Myoblast fusion in Drosophila. BioEssays 24, 591–601.
Gilbert, S. F. (2000) Developmental Biology, 6th ed. Sinauer, Sunderland, MA.
Singer, S. J. (1992) Intercellular communication and cell-cell adhesion. Science 255, 1671–1677.
Dustin, M. L. and Colman, D. R. (2002) Neural and immunological synaptic relations. Science 29, 785–789.
Grakoui, A., Bromley, S. K., Sumen, C., Davis, M. M., Shaw, A. S., Allen, P. M., et al. (1999) The immunological synapse: a molecular machine controlling T-cell activation. Science 285, 221–227.
Weber, I. (2003) Reflection interference contrast microscopy. Meth. Enzymol. 361, 34–47.
Wiegand, G., Neumaier, K. R., and Sackmann, E. (1998) Microinterferometry: three-dimensional reconstruction of surface microtopography for thin-film and wetting studies by reflection interference contrast microscopy (RICM). Appl. Opt. 37, 6892–6905.
Axelrod, D., Hellen, E. H., and Fulbright, R. M. (1992) Total internal reflection fluorescence, in Topics in Fluorescence Spectroscopy, vol. 3 (Lakowicz, J. R., ed.), Plenum, New York.
Thompson, N. L., Drake, A. W., Chen, L., and Vanden Broek, W. (1997) Equilibrium, kinetics, diffusion and self-association of proteins at membrane surfaces: measurement by total internal reflection fluorescence microscopy. Photochem. Photobiol. 65, 39–46.
Ajo-Franklin C. M., Kam, L., and Boxer, S. G. (2001) High refractive index substrates for fluorescence microscopy of biological interfaces with high z contrast. Proc. Natl. Acad. Sci. USA 98, 13643–13648.
Selvin, P. R. (2000) The renaissance of fluorescence resonance energy transfer. Nat. Struct. Biol. 7, 730–734.
Clegg, R. M. (1995) Fluorescence resonance energy transfer. Curr. Op. Biotech. 6, 103–110.
Clegg, R. M. (1996) Fluorescence resonance energy transfer, in Fluorescence Imaging Spectroscopy and Microscopy (Wang, X. F. and Herman, B., ed.), John Wiley & Sons, New York.
Lakowicz, J. R. (1999) Principles of Fluorescence Spectroscopy, Kluwer Academic/Plenum, New York.
Periasami, A. (2001) Fluorescence resonance energy transfer microscopy: a mini-review. J. Biomed. Opt. 6, 287–291.
Wu, P. and Brand, L. (1994) Resonance energy transfer: methods and applications. Anal. Biochem. 218, 1–13.
Förster, T. (1948) Zwischenmolekulare energiewanderung und fluoreszenz. Ann. Physik. 2, 55–75.
Förster, T. (1959) Transfer mechanisms of electronic excitation. Discuss. Faraday Soc. 27, 7–17.
Förster, T. (1965) Delocalized excitation and excitation transfer, in Modern Quantum Chemistry, vol. 3 (Sinanoglu, O., ed.), Academic Press, New York.
Kuhn, H. (1971) Classical aspects of energy transfer in molecular systems. J. Chem. Phys. 53, 101–108.
Stryer, L. and Haugland, R. P. (1967) Energy transfer: a spectroscopic ruler. Proc. Natl. Acad. Sci. USA 58, 719–726.
Stryer, L. (1978) Fluorescence resonance energy transfer as a spectroscopic ruler. Annu. Rev. Biochem. 47, 819–846.
Clegg, R. M., Murchie, A. I. H., Zechel, A., and Lilley, D. M. J. (1993) Observing the helical geometry of double-stranded DNA in solution by fluorescence resonance energy transfer. Proc. Natl. Acad. Sci. USA 90, 2994–2998.
Mergny, J.-L., Boutorine, A. S., Garestier, T., Belloc, G., Rougé, M., Bulychev, N. V., et al. (1994) Fluorescence resonance energy transfer as a probe for nucleic acid structures and sequences. Nucleic Acids Res. 22, 920–928.
Suzuki, Y., Yasunaga, T., Ohkura, R., Wakabayashi, T., and Sutoh, K. (1998) Swing of the lever arm of a myosin motor at the isomerization and phosphate-release steps. Nature 396, 380–383.
Sadqi, M., Lapidus, L. J., and Muñoz, V. (2003) How fast is protein hydrophobic collapse?. Proc. Natl. Acad. Sci. USA 100, 12117–12122.
Corbalan-Garcia, S., Teruel, J. A., and Gomez-Fernandez, J. C. (1993) Intramolecular distances within the Ca2+-ATPase from sarcoplasmic reticulum as estimated through fluorescence energy transfer between probes. Eur. J. Biochem. 217, 737–744.
Baker, K. J., East, J. M., and Lee, A. C. (1994) Localization of the hinge region of the Ca2+-ATPase of sarcoplasmic reticulum using resonance energy transfer. Biochim. Biophys. Acta 1192, 53–60.
Miyawaki, A. and Tsien, R. Y. (2000). Monitoring protein conformations and interactions by fluorescence resonance energy transfer between mutants of green fluorescent protein. Meth. Enzym. 327, 472–500.
Varma, R. and Mayor, S. (1998) GPI-anchored proteins are organized in submicron domains at the cell surface. Nature 394, 798–801.
Kenworthy, A. K., Petranova, N., and Edidin, M. (2000) High-resolution FRET microscopy of cholera toxin B-subunit and GPI-anchored proteins in cell plasma membranes. Mol. Biol. Cell 11, 1645–1655.
Niles, W. D., Silvius, J. R., and Cohen, F. S. (1996) Resonance energy transfer imaging of phospholipid vesicle interaction with a planar phospholipid membrane: undulations and attachment sites in the region of calcium-mediated membrane-membrane adhesion. J. Gen. Physiol. 107, 329–351.
Wong, A. P. and Groves, J. T. (2001) Topographical imaging of an intermembrane junction by combined fluorescence interference and energy transfer microscopies. J. Am. Chem. Soc. 123, 12414–12415.
Wong, A. P. and Groves, J. T. (2002) Molecular topography imaging by intermembrane fluorescence resonance energy transfer. Proc. Natl. Acad. Sci. USA 99, 14147–14152.
Sackmann, E. (1996) Supported membranes: scientific and practical applications. Science 271, 43–48.
Boxer, S. G. (2000) Molecular transport and organization in supported lipid membranes. Curr. Opin. Chem. Biol. 4, 704–709.
Groves, J. T. and Boxer, S. G. (2002) Micropattern formation in supported lipid membranes. Acc. Chem. Res. 35, 149–157.
Merritt, E. A., Sarfaty, S., Akker, F. V. D., L'Hoir, C., Martial, J. A., and Hol, W. G. J. (1994) Crystal structure of cholera toxin B-pentamer bound to receptor G(M1) pentasaccharide. Protein Sci. 3, 166–175.
Parthasarathy, R. and Groves, J. T. (2004) Nonequilibrium adhesion patterns at lipid bilayer junctions. J. Phys. Chem. B. 108, 649–657.
Kaizuka, Y. and Groves, J. T. (2004) Structure and dynamics of supported intermembrane junctions. Biophys. J. 86, 905–912.
Zeck, G. and Fromherz, P. (2003) Repulsion and attraction by extracellular matrix protein in cell adhesion studied with nerve cells and lipid vesicles on silicon chips. Langmuir 19, 1580–1585.
Iwanaga, Y., Braun, D., and Fromherz, P. (2001) No correlation of focal contacts and close adhesion by comparing GFP-vinculin and fluorescence interference of DiI. Eur. Biophys. J. 30, 17–26.
Kiessling, V., and Tamm, L. K. (2003) Measuring distances in supported bilayers by fluorescence interference-contrast microscopy: polymer supports and SNARE proteins. Biophys. J. 84, 408–418.
Lambacher, A. and Fromherz, P. (1996) Fluorescence interference-contrast microscopy on oxidized silicon using a monomolecular dye layer. Appl. Phys. A 63, 207–216.
Braun, D. and Fromherz, P. (1997) Fluorescence interference contrast microscopy of cell adhesion on silicon. Appl. Phys. A 65, 341–348.
Lambacher, A. and Fromherz, P. (2002) Luminescence of dye molecules on oxidized silicon and fluorescence interference contrast microscopy of biomembranes. J. Opt. Soc. Am. B 19, 1435–1453.
Jellison, G. E. and Modine, F.A. (1982) Optical constants for silicon at 300 and 10 K determined from 1.64 to 4.73 eV by ellipsometry. J. Appl. Phys. 53, 3745–3753.
Landolt, H. and Börnstein, R. (1962). Numerical Data and Functional Relationships in Science and Technology, 6th ed., Vol. 2, Springer, Berlin.
Swan, A. K., Moiseev, L. A., Cantor, C. R., Davis, B., Ippolito, S. B., Karl, W. C., et al. (2003) Toward nanometer-scale resolution in fluorescence microscopy using spectral self-interference. IEEE J. Sel. Top. Quantum Electron. 9, 294–300.
Eah, S.-K., Jaeger, H. M., Scherer, N. F., Wiederrecht, G. P., and Lin, X.-M. (manuscript in preparation).
Medhage, B., Mukhtar, E., Kalman, B., Johansson, L., and Molotkovsky, J. G. (1992) J. Chem. Soc. Faraday Trans. 88, 2845–2841.
Nardi, J., Bruinsma, R., and Sackmann, E. (1998) Adhesion-induced reorganization of charged fluid membranes. Phys. Rev. E 58, 6340–6354.
Gu, M. (2000) Advanced Optical Imaging Theory, Springer, Heidelberg.
Parthasarathy, R. and Groves, J. T. (2004) Protein patterns at lipid bilayer junctions. Proc. Natl. Acad. Sci. USA 101, 12,798–12,803.
Curtis, A. S. G. (1964) The mechanism of adhesion of cells to glass. A study by interference reflection microscopy. J. Cell Biol. 20, 199–215.
Ploem, J. S. (1975). Reflection contrast microscopy as a tool for investigation of the attachment of living cells to a glass surface, in Mononuclear Phagocytes in Immunity Infection and Pathology (van Furth, R., ed.), Blackwell, Oxford.
Rädler, J. and Sackmann, E. (1993) Imaging optical thicknesses and separation distances of phospholipid vesicles at solid surfaces. J. Phys. II France 3, 727–748.
Verschueren, H. (1985) Interference reflection microscopy in cell biology: methodology and applications. J. Cell. Sci. 75, 279–301.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Parthasarathy, R., Groves, J.T. Optical techniques for imaging membrane topography. Cell Biochem Biophys 41, 391–414 (2004). https://doi.org/10.1385/CBB:41:3:391
Issue Date:
DOI: https://doi.org/10.1385/CBB:41:3:391