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High-Resolution Analysis of the 3D Organization of Human Metaphase Chromosomes

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Atomic Force Microscopy

Part of the book series: Methods in Molecular Biology™ ((MIMB,volume 242))

Abstract

The detailed understanding of the nuclear cell functions requires an accurate knowledge of the spatial organization of their structures. For many years the study of human metaphase chromosomes was conducted with light microscopy after staining protocols disturbing the native chromosomal structure. The approach by scanning electron microscopy (SEM) provides higher resolution as compared with light microscopy and permits surface analysis of the chromosomal structure, which cannot be adequately obtained from transmission electron microscopy. Nevertheless, to obtain high resolution in SEM observations, the use of a high electron accelerating voltage (up to 30 kV) is required (1). With these experimental conditions, the sputter coating or a conductive staining of the samples is generally required (1,2). Both procedures allow electron-charging dispersion from the sample but may obscure fine details and produce sample alterations (3).

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References

  1. Sumner, A. T., Ross, A. R., and Graham, E. (1994) Preparation of chromosomes for scanning electron microscopy. Methods Mol. Biol. 29, 41–50.

    PubMed  CAS  Google Scholar 

  2. Wanner, G. and Formanek, H. (1995) Imaging of DNA in human and plant chromosomes by high-resolution scanning electron microscopy. Chromosome Res. 3, 368–374.

    Article  PubMed  CAS  Google Scholar 

  3. Hermann, R. and Müller, M. (1992) Towards high resolution SEM of biological objects. Arch. Histol. Cytol. 55, 17–25.

    Article  PubMed  Google Scholar 

  4. Nagatani, T., Saito, S., Sato, M., and Yamada, M. (1987) Development of an ultra high resolution scanning electron microscope by means of a field emission source and in-lens system. Scanning Microsc. 1, 901–909.

    Google Scholar 

  5. Pawley, J. (1997) The development of field-emission scanning electron microscopy for imaging biological surfaces. Scanning 19, 324–336.

    PubMed  CAS  Google Scholar 

  6. Rizzoli, R., Rizzi, E., Falconi, M., Galanzi A., Baratta B., Lattanzi, G., et al. (1994) High resolution detection of uncoated metaphase chromosome by means of field emission scanning electron microscopy. Chromosoma 103, 393–400.

    Article  PubMed  CAS  Google Scholar 

  7. E., Falconi, M., Baratta, B., Manzoli, L., Galanzi, A., Lattanzi, G., et al. (1995) High-resolution FEISEM detection of DNA centromeric probes in HeLa metaphase chromosomes. J. Histochem. Cytochem. 43, 413–419.

    PubMed  Google Scholar 

  8. Lattanzi, G., Galanzi, A., Gobbi, P., Falconi, M., Matteucci, A., Breschi, L., et al. (1998) Ultrastructural aspects of the DNA polymerase distribution during the cell cycle. J. Histochem. Cytochem. 46, 1435–1442.

    PubMed  CAS  Google Scholar 

  9. Gobbi, P., Falconi, M., Vitale, M., Galanzi, A., Artico, M., Martelli, A. M., et al. (1999) Scanning electron microscopic detection of nuclear structures involved in DNA replication. Arch. Histol. Cytol. 62, 317–326.

    Article  PubMed  CAS  Google Scholar 

  10. Binnig, G., Quate, C. F., and, Gerber, C. H. (1986) Atomic force microscopy. Phys. Rev. Lett. 56, 930–933.

    Article  PubMed  Google Scholar 

  11. Mariani, T., Musio, A., Frediani, C., Sbrana, I., and Ascoli, C. (1994) An atomic force microscope for cytological and histological investigations. J. Microsc. 176, 121–131.

    PubMed  CAS  Google Scholar 

  12. Ushiki, T., Hitomi, J., Ogura, S., Umemoto, T., and Shigeno, M. (1996) Atomic force microscopy in hystology and cytology. Arch. Histol. Cytol. 50, 421–431.

    Google Scholar 

  13. Putman, C. A.J., Van der Werf, K. O., De Grooth, B. G., Van Hulst, N. F., Segerink, F. B. and Greve, J. (1992) Atomic force microscope featuring an integrated optical microscope. Ultramicroscopy 42/44, 1549–1552.

    Article  Google Scholar 

  14. Musio, A., Mariani, T., Frediani, C., Ascoli, C., and Sbrana, I. (1997) Atomic force microscope imaging of chromosome structure during G-banding treatments. Genome 40, 127–131.

    Article  PubMed  CAS  Google Scholar 

  15. Thalhammer, S., Köhler, U., Stark, R., and Heckl, W. M. (2000) GTG banding pattern on human metaphase chromosomes revealed by high resolution atomic-force microscopy. J. Microsc. 202, 464–467.

    Article  Google Scholar 

  16. Gobbi, P., Thalhammer, S., Falconi, M., Stark, R., Heckl, W. M., Mazzotti, G. (2000) Correlative high resolution morphological analysis of the three-dimensional organization of human metaphasechromosomes. Scanning 22, 273–281.

    Article  PubMed  CAS  Google Scholar 

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© 2004 Humana Press Inc., Totowa, NJ

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Thalhammer, S., Gobbi, P., Falconi, M., Mazzotti, G., Heckl, W.M. (2004). High-Resolution Analysis of the 3D Organization of Human Metaphase Chromosomes. In: Braga, P.C., Ricci, D. (eds) Atomic Force Microscopy. Methods in Molecular Biology™, vol 242. Humana Press. https://doi.org/10.1385/1-59259-647-9:245

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  • DOI: https://doi.org/10.1385/1-59259-647-9:245

  • Publisher Name: Humana Press

  • Print ISBN: 978-1-58829-094-6

  • Online ISBN: 978-1-59259-647-8

  • eBook Packages: Springer Protocols

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