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. 1999 Jan;76(1 Pt 1):509–516. doi: 10.1016/s0006-3495(99)77219-x

Imaging of cell/substrate contacts of living cells with surface plasmon resonance microscopy.

K Giebel 1, C Bechinger 1, S Herminghaus 1, M Riedel 1, P Leiderer 1, U Weiland 1, M Bastmeyer 1
PMCID: PMC1302541  PMID: 9876164

Abstract

We have developed a new method for observing cell/substrate contacts of living cells in culture based on the optical excitation of surface plasmons. Surface plasmons are quanta of an electromagnetic wave that travel along the interface between a metal and a dielectric layer. The evanescent field associated with this excitation decays exponentially perpendicular to the interface, on the order of some hundreds of nanometers. Cells were cultured on an aluminum-coated glass prism and illuminated from below with a laser beam. Because the cells interfere with the evanescent field, the intensity of the reflected light, which is projected onto a camera chip, correlates with the cell/substrate distance. Contacts between the cell membrane and the substrate can thus be visualized at high contrast with a vertical resolution in the nanometer range. The lateral resolution along the propagation direction of surface plasmons is given by their lateral momentum, whereas perpendicular to it, the resolution is determined by the optical diffraction limit. For quantitative analysis of cell/substrate distances, cells were imaged at various angles of incidence to obtain locally resolved resonance curves. By comparing our experimental data with theoretical surface plasmon curves we obtained a cell/substrate distance of 160 +/- 10 nm for most parts of the cells. Peripheral lamellipodia, in contrast, formed contacts with a cell substrate/distance of 25 +/- 10 nm.

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Selected References

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  1. Axelrod D., Thompson N. L., Burghardt T. P. Total internal inflection fluorescent microscopy. J Microsc. 1983 Jan;129(Pt 1):19–28. doi: 10.1111/j.1365-2818.1983.tb04158.x. [DOI] [PubMed] [Google Scholar]
  2. Bastmeyer M., Bähr M., Stuermer C. A. Fish optic nerve oligodendrocytes support axonal regeneration of fish and mammalian retinal ganglion cells. Glia. 1993 May;8(1):1–11. doi: 10.1002/glia.440080102. [DOI] [PubMed] [Google Scholar]
  3. Bastmeyer M., Jeserich G., Stuermer C. A. Similarities and differences between fish oligodendrocytes and Schwann cells in vitro. Glia. 1994 Aug;11(4):300–314. doi: 10.1002/glia.440110403. [DOI] [PubMed] [Google Scholar]
  4. Burmeister J. S., Olivier L. A., Reichert W. M., Truskey G. A. Application of total internal reflection fluorescence microscopy to study cell adhesion to biomaterials. Biomaterials. 1998 Mar;19(4-5):307–325. doi: 10.1016/s0142-9612(97)00109-9. [DOI] [PubMed] [Google Scholar]
  5. Burmeister J. S., Truskey G. A., Reichert W. M. Quantitative analysis of variable-angle total internal reflection fluorescence microscopy (VA-TIRFM) of cell/substrate contacts. J Microsc. 1994 Jan;173(Pt 1):39–51. doi: 10.1111/j.1365-2818.1994.tb03426.x. [DOI] [PubMed] [Google Scholar]
  6. CURTIS A. S. THE MECHANISM OF ADHESION OF CELLS TO GLASS. A STUDY BY INTERFERENCE REFLECTION MICROSCOPY. J Cell Biol. 1964 Feb;20:199–215. doi: 10.1083/jcb.20.2.199. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Davies P. F., Robotewskyj A., Griem M. L. Endothelial cell adhesion in real time. Measurements in vitro by tandem scanning confocal image analysis. J Clin Invest. 1993 Jun;91(6):2640–2652. doi: 10.1172/JCI116503. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Davies P. F., Robotewskyj A., Griem M. L. Quantitative studies of endothelial cell adhesion. Directional remodeling of focal adhesion sites in response to flow forces. J Clin Invest. 1994 May;93(5):2031–2038. doi: 10.1172/JCI117197. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Gingell D. The interpretation of interference-reflection images of spread cells: significant contributions from thin peripheral cytoplasm. J Cell Sci. 1981 Jun;49:237–247. doi: 10.1242/jcs.49.1.237. [DOI] [PubMed] [Google Scholar]
  10. Izzard C. S., Lochner L. R. Cell-to-substrate contacts in living fibroblasts: an interference reflexion study with an evaluation of the technique. J Cell Sci. 1976 Jun;21(1):129–159. doi: 10.1242/jcs.21.1.129. [DOI] [PubMed] [Google Scholar]
  11. Kolega J., Shure M. S., Chen W. T., Young N. D. Rapid cellular translocation is related to close contacts formed between various cultured cells and their substrata. J Cell Sci. 1982 Apr;54:23–34. doi: 10.1242/jcs.54.1.23. [DOI] [PubMed] [Google Scholar]
  12. Lanni F., Waggoner A. S., Taylor D. L. Structural organization of interphase 3T3 fibroblasts studied by total internal reflection fluorescence microscopy. J Cell Biol. 1985 Apr;100(4):1091–1102. doi: 10.1083/jcb.100.4.1091. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Mrksich M., Whitesides G. M. Using self-assembled monolayers to understand the interactions of man-made surfaces with proteins and cells. Annu Rev Biophys Biomol Struct. 1996;25:55–78. doi: 10.1146/annurev.bb.25.060196.000415. [DOI] [PubMed] [Google Scholar]
  14. Paddock S. W. Tandem scanning reflected-light microscopy of cell-substratum adhesions and stress fibres in Swiss 3T3 cells. J Cell Sci. 1989 May;93(Pt 1):143–146. doi: 10.1242/jcs.93.1.143. [DOI] [PubMed] [Google Scholar]
  15. Panayotou G., Waterfield M. D., End P. Riding the evanescent wave. Curr Biol. 1993 Dec 1;3(12):913–915. doi: 10.1016/0960-9822(93)90236-h. [DOI] [PubMed] [Google Scholar]
  16. Verschueren H. Interference reflection microscopy in cell biology: methodology and applications. J Cell Sci. 1985 Apr;75:279–301. doi: 10.1242/jcs.75.1.279. [DOI] [PubMed] [Google Scholar]

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