A second generation apparatus for time-resolved electron cryo-microscopy using stepper motors and electrospray

A second generation apparatus for time-resolved electron cryo-microscopy using stepper motors and... We describe here a second generation apparatus for studying transient reaction conformations in macromolecules and their complexes by electron cryo-microscopy. Reactions are trapped by rapid freezing in times ranging from a few milliseconds to tens of seconds after initiation. Blotting of the electron microscope grid and freezing it in liquid ethane uses computer controlled microstepping motors. For the fastest time resolution, a blotted grid containing a thin film of one reactant is sprayed with small droplets containing a second reactant just before freezing. The spray is produced electrically (electrospray), which gives a dense cloud of droplets <1 μm in diameter from the 1–2 μl of solution required per grid. A second method in which two solutions are first mixed by turbulent flow and then blotted prior to freezing is used for reactions with time courses >1 s. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Journal of Structural Biology Elsevier

A second generation apparatus for time-resolved electron cryo-microscopy using stepper motors and electrospray

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Publisher
Elsevier
Copyright
Copyright © 2003 Elsevier Inc.
ISSN
1047-8477
eISSN
1095-8657
DOI
10.1016/j.jsb.2003.09.027
Publisher site
See Article on Publisher Site

Abstract

We describe here a second generation apparatus for studying transient reaction conformations in macromolecules and their complexes by electron cryo-microscopy. Reactions are trapped by rapid freezing in times ranging from a few milliseconds to tens of seconds after initiation. Blotting of the electron microscope grid and freezing it in liquid ethane uses computer controlled microstepping motors. For the fastest time resolution, a blotted grid containing a thin film of one reactant is sprayed with small droplets containing a second reactant just before freezing. The spray is produced electrically (electrospray), which gives a dense cloud of droplets <1 μm in diameter from the 1–2 μl of solution required per grid. A second method in which two solutions are first mixed by turbulent flow and then blotted prior to freezing is used for reactions with time courses >1 s.

Journal

Journal of Structural BiologyElsevier

Published: Oct 1, 2003

References

  • Some tenets pertaining to electrospray ionization mass spectrometry
    Cole, R.B.
  • A brief overview of the present status of the mechanisms involved in electrospray mass spectrometry
    Kebarle, P.
  • Freeze trapping of reaction intermediates
    Moffat, K.; Henderson, R.

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