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J Cell Biol. May 1, 1973; 57(2): 551–565.
PMCID: PMC2108989

A TECHNIQUE FOR ULTRACRYOTOMY OF CELL SUSPENSIONS AND TISSUES

Abstract

Ultracryotomy of fixed tissue has been investigated for a number of years but, so far, success has been limited for several reasons. The simple technique herein reported allows the ultracryotomy not only of a variety of tissues but also of single cells in suspension, with a preservation and visualization of ultrastructural detail at least equivalent to that obtained with conventional embedding procedures. In this technique, sucrose is infused into glutaraldehyde-fixed tissue pieces before freezing for the purpose of controlling the sectioning consistency. By choosing the proper combinations of sucrose concentration and sectioning temperature, a wide variety of tissues can be smoothly sectioned. Isolated cells, suspended in a sucrose solution, are sectioned by sectioning the frozen droplet of the suspension. A small liquid droplet of a saturated or near-saturated sucrose solution, suspended on the tip of an eyelash probe, is used to transfer frozen sections from the knife edge onto a grid substrate or a water surface. Upon melting of the sections on the surface of the sucrose droplet, they are spread flat and smooth due to surface tension. When the section of a suspension of single cells melts, individual sections of cells remain confined to the small area of the droplet surface. These devices make it possible to cut wide dry sections, and to avoid flotation on dimethyl sulfoxide solutions. With appropriate staining procedures, well-preserved ultrastructural detail can be observed. The technique is illustrated with a number of tissue preparations and with suspensions of erythrocytes and bacterial cells.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Bernhard W, Leduc EH. Ultrathin frozen sections. I. Methods and ultrastructural preservation. J Cell Biol. 1967 Sep;34(3):757–771. [PMC free article] [PubMed]
  • Bernhard W, Viron A. Improved techniques for the preparation of ultrathin frozen sections. J Cell Biol. 1971 Jun;49(3):731–746. [PMC free article] [PubMed]
  • Iglesias JR, Bernier R, Simard R. Ultracryotomy: a routine procedure. J Ultrastruct Res. 1971 Jul;36(1):271–289. [PubMed]
  • Bernier R, Iglesias R, Simard R. Detection of DNA by tritiated actinomycin D on ultrathin frozen sections. J Cell Biol. 1972 Jun;53(3):798–808. [PMC free article] [PubMed]
  • McLean JD, Singer SJ. A general method for the specific staining of intracellular antigens with ferritin-antibody conjugates. Proc Natl Acad Sci U S A. 1970 Jan;65(1):122–128. [PMC free article] [PubMed]
  • Christensen AK. Frozen thin sections of fresh tissue for electron microscopy, with a description of pancreas and liver. J Cell Biol. 1971 Dec;51(3):772–804. [PMC free article] [PubMed]
  • MARINOZZI V. CYTOCHIMIE ULTRASTRUCTURALE DU NUCL'EOLE--RNA ET PROT'EINES INTRANUCL'EOLAIRES. J Ultrastruct Res. 1964 Jun;10:433–456. [PubMed]
  • Nicolson GL. Structure of the photosynthetic apparatus in protein-embedded chloroplasts. J Cell Biol. 1971 Jul;50(1):258–263. [PMC free article] [PubMed]
  • Caspar DL, Kirschner DA. Myelin membrane structure at 10 A resolution. Nat New Biol. 1971 May 12;231(19):46–52. [PubMed]
  • Knappeis GG, Carlsen F. The ultrastructure of the M line in skeletal muscle. J Cell Biol. 1968 Jul;38(1):202–211. [PMC free article] [PubMed]
  • Rowe RW. Ultrastructure of the Z line of skeletal muscle fibers. J Cell Biol. 1971 Dec;51(3):674–685. [PMC free article] [PubMed]
  • HUXLEY HE. The double array of filaments in cross-striated muscle. J Biophys Biochem Cytol. 1957 Sep 25;3(5):631–648. [PMC free article] [PubMed]
  • FARQUHAR MG, PALADE GE. Segregation of ferritin in glomerular protein absorption droplets. J Biophys Biochem Cytol. 1960 Apr;7:297–304. [PMC free article] [PubMed]

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