[3dem] amplitude contrast valuesHi,

[Masahide Kikkawa]

Hi David,

I would follow this procedure and test using your own specimen:

Ultramicroscopy. 1988;25(4):279-91.
Contrast transfer for frozen-hydrated specimens: determination from pairs of defocused images.
Toyoshima C, Unwin N.

Abstract
Electron imaging of frozen-hydrated biological molecules allows density maps to be obtained directly, without the need for fixatives or stains. The appearance of such maps may, however, be strongly influenced by the contrast transfer properties, which have not previously been evaluated by quantitative experiments. Here we determine the contribution due to amplitude contrast in a typical (approximately 300 A thick) frozen specimen, consisting of arrays of acetylcholine receptor, by comparing pairs of images recorded with different defocuses. We find that this specimen is imaged as a "weak-phase-weak-amplitude" object and that the contribution due to amplitude contrast is 7%.


On Jul 22, 2014, at 2:02 AM, David Gene Morgan <dagmorga at indiana.edu> wrote:

> Hi,
> 
> 	I'm trying to do some automated defocus determination (initially trying Niko's ctffind3 program) and would like some input on appropriate amplitude contrast values for various types of samples.  I know that ice embedded images usually are said to have an amplitude contrast of 0.07 to 0.1.  I also know that with protein/nucleic acid complexes in ice, amplitude contrast may need to be set a bit higher (say to as much as 0.15 for some data I worked on awhile ago).
> 
> 	But I don't have any sort of feeling for appropriate values of amp;itude contrast for different sorts of negative stains (either cryo negative stain or standard negative stain).  It should be "high," but I don't know how high might be appropriate.
> 
> 	If anyone has either some practical or theoretical insight into this, I'd appreciate some suggestions.  Also, does anyone know about an appropriate amplitude contrast for a standard Au/Pd waffle grid?
> 
> 	Thanks in advance for all your help.
> 
> -- 
>                 David Gene Morgan
>             Electron Microscopy Center
>                  047D Simon Hall
>                  IU Bloomington
>               812 856 1457 (office)
>               812 856 3221 (3200)
>            http://bio.indiana.edu/~cryo
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---------------------------------------------------------------
Masahide Kikkawa, M.D., Ph. D.
Professor
Department of Cell Biology & Anatomy
Graduate School of Medicine
The University of Tokyo
http://structure.m.u-tokyo.ac.jp/
tel: +81-3-5841-3338
fax: +81-3-5841-3339
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