Atomic force microscopy of native human metaphase chromosomes in a liquid

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  • Hoshi Osamu
    Division of Microscopic Anatomy and Bio-imaging, Niigata University Graduate School of Medical and Dental Sciences
  • Shigeno Masatsugu
    SII NanoTechnology Inc.
  • Ushiki Tatsuo
    Division of Microscopic Anatomy and Bio-imaging, Niigata University Graduate School of Medical and Dental Sciences

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Abstract

The present study introduces a method for obtaining three-dimensional images of native (i.e., unfixed) chromosomes by atomic force microscopy (AFM) in a liquid. Human metaphase chromosomes were isolated from a human lymphoblast-like cell line, K562, by the hexylene glycol procedure according to Wray and Stubble- field (1970), adsorbed on a silane-coated glass slide, and observed in a dynamic force mode (i.e., intermittent contact mode) of AFM in a hexylene buffer solution. In adequate operating conditions, the shape of chromosomes with paired chromatids was clearly and three-dimensionally observed by AFM. At high magnification, globular or fibrous structures about 50 nm thick could be found on the surface of each chromaid, implying that chromatin fibers were strongly wound or twisted in the chromatid. Thus, AFM imaging enabled the direct visualization of native chromosomes in a liquid at high resolution - which is com parable with that of scanning electron microscopy - and can serve to analyze the mechanism of chromosome condensation and separation in relation to the structure of chromosomes.

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