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Mixing Tandem Cryo-Hrsem and Cryo-Stem Images of Cells and Macromolecular Assembles

Published online by Cambridge University Press:  02 July 2020

R.P. Apkarian
R.P. Apkarian*
Affiliation:
Integrated Microscopy & Microanalytical Facility, Department of Chemistry, Emory University, Atlanta, GA, 30322
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Abstract

Creating contrast of unstained frozen hydrated cells and isolated macromolecules in order to provide high fidelity recordings present several difficulties and therefor require attempts to maximize image information for characterization purposes. Adobe Photoshop 5.0 was employed to overlay (“layer“) topographic high resolution secondary electron (SE-I) images (HRSEM) onto STEM images with and without polarity reversal employed during digital image acquisition. Cells and molecular samples precipitated from an aqueous droplet or aerosol and supported on carbon and holey carbon coated grids were plunge frozen and Chromium (Cr) coated. Since Cr-coated cryosamples on carbon-grid supports have very low mass density and because signal to noise is low in HRSEM recordings, especially in regions which overhang holey carbon expanses, then the combined contrasts obtained in a composite image containing STEM and HRSEM contrasts result in greater detail recognition.

Tandem cryo-imaging, using an in-lens field emission (FE) SEM (DS-130F) operated at 25kV and fitted with above-lens secondary electron detector and below-lens scanning transmission detector, provided digital images of frozen cells and macromolecular complexes.

Type
Cryoimmobilization, Freeze Substitution and Cryoem (Organized by S. Erlandsen)
Information
Microscopy and Microanalysis , Volume 7 , Issue S2: Proceedings: Microscopy & Microanalysis 2001, Microscopy Society of America 59th Annual Meeting, Microbeam Analysis Society 35th Annual Meeting, Long Beach, California August 5-9, 2001 , August 2001 , pp. 722 - 723
Copyright
Copyright © Microscopy Society of America 2001

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References

References:

1.Apkarian, R.P. et al., Microsc. & Microanal. 5 (1999)197.CrossRefGoogle Scholar
2.Apkarian, R.P. et al., Proc. Ann. Mtg MSA (2000) 292.CrossRefGoogle Scholar
3.Lee, et al., Adv. Mater. (2000) 12:15; 1105.3.0.CO;2-1>CrossRefGoogle Scholar
4.Ms.Wright, E.R., Ms.Zhou, .J. and Dr.Conticello, V.P. are acknowledged for their technical assistance and for providing protein specimens.Google Scholar