title: Staining and embedding of human chromosomes for 3-d serial block-face scanning electron microscopy.
abstract: The high-order structure of human chromosomes is an important biological question that is still under investigation. Studies have been done on imaging human mitotic chromosomes using mostly 2-D microscopy methods. To image micron-sized human chromosomes in 3-D, we developed a procedure for preparing samples for serial block-face scanning electron microscopy (SBFSEM). Polyamine chromosomes are first separated using a simple filtration method and then stained with heavy metal. We show that the DNA-specific platinum blue provides higher contrast than osmium tetroxide. A two-step procedure for embedding chromosomes in resin is then used to concentrate the chromosome samples. After stacking the SBFSEM images, a familiar X-shaped chromosome was observed in 3-D.
authors.affiliation: London Centre for Nanotechnology, University College London, London, UK; Research Complex at Harwell, Rutherford Appleton Laboratory, Oxon, UK.
authors.forename: Mohammed
authors.initials: M
authors.lastname: Yusuf
authors.affiliation: London Centre for Nanotechnology, University College London, London, UK; Research Complex at Harwell, Rutherford Appleton Laboratory, Oxon, UK.
authors.forename: Bo
authors.initials: B
authors.lastname: Chen
authors.affiliation: Corrosion & Protection Centre, School of Materials, The University of Manchester, Manchester, UK.
authors.forename: Teruo
authors.initials: T
authors.lastname: Hashimoto
authors.affiliation: London Centre for Nanotechnology, University College London, London, UK; Research Complex at Harwell, Rutherford Appleton Laboratory, Oxon, UK.
authors.forename: Ana Katrina
authors.initials: AK
authors.lastname: Estandarte
authors.affiliation: Corrosion & Protection Centre, School of Materials, The University of Manchester, Manchester, UK.
authors.forename: George
authors.initials: G
authors.lastname: Thompson
authors.affiliation: London Centre for Nanotechnology, University College London, London, UK; Research Complex at Harwell, Rutherford Appleton Laboratory, Oxon, UK.
authors.forename: Ian
authors.initials: I
authors.lastname: Robinson
mesh_terms: D002460:Cell Line; D002877:Chromosomes, Human / Q000737:chemistry*; D004396:Coloring Agents / Q000737:chemistry; D016591:Histocytological Preparation Techniques / Q000379:methods*; D006801:Humans; D008297:Male; D008855:Microscopy, Electron, Scanning / Q000379:methods*; D008938:Mitosis; D010984:Platinum / Q000737:chemistry
chemical_list: D004396:Coloring Agents; D010984:Platinum
keywords: chromosome; heavy metal staining; imaging; platinum blue; serial block-face electron microscopy; three-dimensional