A Perspective on Biological X-Ray and Electron Microscopy

  • Chris Jacobsen
  • Robin Medenwaldt
  • Shawn Williams


We consider image contrast for electron microscopy of thick hydrated biological specimens, allowing for the use of phase contrast and energy filtering. This allows us to gain perspective on the relative roles of electron and soft X-ray microscopes. Radiation dose is found to depend strongly on ice thickness, with electrons offering lower dose if the ice thickness is less than about 500 nm, and x rays offering lower dose for thicker ice layers.


Elastic Scattering Inelastic Scattering Electron Energy Loss Spectroscopy Objective Aperture Energy Filter 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


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  1. 1.
    E. Betzig, J. K. Trautman, T. D. Harris, J. S. Weiner, and R. L. Kostelak. Breaking the diffraction barrier: optical microscopy on a nanometric scale. Science, 251:1468–1470,1991.ADSCrossRefGoogle Scholar
  2. 2.
    W. A. Carrington, R. M. Lynch, E. D. W. Moore, G. Isenberg, K. E. Fogarty, and F. S. Fay. Superresolution three-dimensional images of fluorescence in cells with minimal light exposure. Science, 268:1483–1487, 1995.ADSCrossRefGoogle Scholar
  3. 3.
    J. R. Breedlove, Jr. and G. T. Trammel. Molecular microscopy: fundamental limitations. Science, 170: 1310–1313, 1970.ADSCrossRefGoogle Scholar
  4. 4.
    R. Henderson. The potential and limitations of neutrons, electrons and X-rays for atomic resolution microscopy of unstained biological molecules. Quarterly Reviews of Biophysics, 28(2):171–193,1995.CrossRefGoogle Scholar
  5. 5.
    D. Sayre, J. Kirz, R. Feder, D. M. Kim, and E. Spiller. Transmission microscopy of unmodified biological materials: Comparative radiation dosages with electrons and ultrasoft x-ray photons. Ultramicroscopy, 2:337–341, 1977.CrossRefGoogle Scholar
  6. 6.
    M. Isaacson and M. Utlat. A comparison of electron and photon beams for determining micro-chemical environment. Optik, 50:213–234, 1978.Google Scholar
  7. 7.
    P. Golz. Calculations on radiation dosages of biological materials in phase contrast and amplitude contrast x-ray microscopy. In A. G. Michette, G. R. Morrison, and C. J. Buckley, editors, X-ray Microscopy III, volume 67 of Springer Series in Optical Sciences, pages 313–315, Berlin, 1992. Springer-Verlag.CrossRefGoogle Scholar
  8. 8.
    R. R. Schroder. Zero-loss energy-filtered imaging of frozen-hydrated proteins: model calculations and implications for future developments. Journal of Microscopy, 166:389–400, 1992.CrossRefGoogle Scholar
  9. 9.
    J. P. Langmore and M. F. Smith. Quantitatitve energy-filtered electron microscopy of biological molecules in ice. Ultramicroscopy, 46:349–373, 1992.CrossRefGoogle Scholar
  10. 10.
    J. P. Langmore, J. Wall, and M.S. Isaacson. The collection of scattered electrons in dark field electron microscopy: I. elastic scattering. Optik, 38:335–350, 1973.Google Scholar
  11. 11.
    J. Wall, M. Isaacson, and J.P. Langmore. The collection of scattered electrons in dark field electron microscopy: II. inelastic scattering. Optik, 39:359–374, 1974. of Emin should be 2.Google Scholar
  12. 12.
    R. A. London, M. D. Rosen, and J. E. Trebes. Wavelength choice for soft x-ray laser holography of biological samples. Applied Optics, 28:3397–3404, 1989.ADSCrossRefGoogle Scholar
  13. 13.
    C. Dinges, A. Berger, and H. Rose. Simulation of TEM images considering phonon and electronic excitations. Ultramicroscopy, 60:49–70, 1995.CrossRefGoogle Scholar
  14. 14.
    S. Q. Sun, S. L. Shi, and R. D. Leapman. Water distribution of hydrated biological specimens by valence electron energy loss spectroscopy. Ultramicroscopy, 50:127-139, 1993.CrossRefGoogle Scholar
  15. 15.
    R. Grimm, D. Typke, M. Barmann, and W. Baumeister. Determination of the inelastic mean free path in ice by examination of tilted vesicles and automated most probable loss imaging. Ultramicroscopy, 63: 169–179, 1996.CrossRefGoogle Scholar
  16. 16.
    M. Isaacson. Inelastic scattering and beam damage of biological molecules. In B. M. Siegel and D. R. Beaman, editors, Physical aspects of electron microscopy and micro beam analysis, pages 247–258, New York, 1975. Wiley.Google Scholar
  17. 17.
    M. J. Berger and S. M. Seltzer. Tables of energy-losses and ranges of electrons and positrons. Technical Report Publication 1133, Committee on Nuclear Science, National Research Council, National Academy of Sciences, Washington, D.C., 1964. Chapter 10, pp. 205–268, Library of Congress catalogue number 64–60027.Google Scholar
  18. 18.
    M. Isaacson, 1994. Personal communication.Google Scholar
  19. 19.
    L. Reimer. I′ransmission electron microscopy: physics of image formation and microanalysis. Springer-Verlag, Berlin, third edition, 1993. Springer Series in Optical Sciences 36.CrossRefGoogle Scholar
  20. 20.
    A. V. Crewe and T. Groves. Thick specimens in the CEM and STEM. I: Contrast. Journal of Applied Physics, 45:3662–3672, 1974.ADSCrossRefGoogle Scholar
  21. 21.
    R. R. Schroder, W. Hofmann, and J.-F. Menetret. Zero-loss energy filtering as improved imaging mode in cryoelectronmicroscopy of frozen-hydrated specimens. Journal of Structural Biology, 105:28–34, 1990.CrossRefGoogle Scholar
  22. 22.
    R. M. Glaeser. Limitations to significant information in biological electron microscopy as a result of radiation damage. Journal of Ultrastructure Research, 36:466–482, 1971.CrossRefGoogle Scholar
  23. 23.
    C. Jacobsen, S. Lindaas, S. Williams, and X. Zhang. Scanning luminescence x-ray microscopy: imaging fluorescence dyes at suboptical resolution. J. Microscopy, 172:121–129, 1993.CrossRefGoogle Scholar
  24. 24.
    M.M. Moronne, C. Larabell, P.R. Selvin, and A. Irtel von Brenndorff. Development offluroescent probes for x-ray microscopy. In G. W. Bailey and A. J. GarrattReed, editors, Proceedings of the 52nd Annual Meeting of the Microscopy Society of America, pages 48–49, San Francisco, 1994. San Francisco Press.Google Scholar
  25. 25.
    H. N. Chapman, J. Fu, C. Jacobsen, and S. Williams. Dark-field x-ray microscopy of immunogold-labeled cells. Journal of the Microscopy Society of America, 2(2):53–62, 1996.Google Scholar
  26. 26.
    R. D. Leapman and S. Sun. Cryo-electron energy loss spectroscopy: observations on vitrified hydrated specimens and radiation damage. Ultramicroscopy, 59:71–79, 1995.CrossRefGoogle Scholar
  27. 27.
    G. Schneider, B. Niemann, P. Guttmann, D. Rudolph, and G. Schmahl. Cryo xray microscopy. Synchrotron Radiation News, 8(3):19–28, 1995.CrossRefGoogle Scholar
  28. 28.
    J. F. Conway, B. L. Trus, F. P. Booy, W. W. Newcomb, J. C. Brown, and A. C. Steven. The effects of radiation damage on the structure of frozen hydrated HSV-1 capsids. Journal of Structural Biology, 111:222–233, 1993.CrossRefGoogle Scholar
  29. 29.
    M. K. Lamvik. Radiation damage in dry and frozen hydrated organic material. Journal of Microscopy, 161:171–181, 1991.CrossRefGoogle Scholar
  30. 30.
    M. F. Schmid, J. Jakana, P. Matsudaira, and W. Chu. Effects of radiation damage with 400-kV electrons on frozen, hydrated actin bundles. Journal of Structural Biology, 108:62–68, 1992.CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 1998

Authors and Affiliations

  • Chris Jacobsen
    • 1
  • Robin Medenwaldt
    • 2
  • Shawn Williams
    • 3
  1. 1.Department of PhysicsSUNY Stony BrookStony BrookUSA
  2. 2.Institute for Storage Ring FacilitiesUniversity of AarhusAarhus CDenmark
  3. 3.Boyer Center for Molecular MedicineYale UniversityNew HavenUSA

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