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Improved Microchip Design and Application for In Situ Transmission Electron Microscopy of Macromolecules

Published online by Cambridge University Press:  13 December 2013

Madeline J Dukes
Affiliation:
Protochips Inc., Raleigh, NC 27606, USA
Rebecca Thomas
Affiliation:
Protochips Inc., Raleigh, NC 27606, USA
John Damiano
Affiliation:
Protochips Inc., Raleigh, NC 27606, USA
Kate L Klein
Affiliation:
Department of Mechanical Engineering, University of the District of Columbia, Washington, DC 20008, USA
Sharavanan Balasubramaniam
Affiliation:
Macromolecules and Interfaces Institute, Virginia Tech, Blacksburg, VA 24061, USA
Sanem Kayandan
Affiliation:
Macromolecules and Interfaces Institute, Virginia Tech, Blacksburg, VA 24061, USA Department of Chemistry, Virginia Tech, Blacksburg, VA 24061, USA
Judy S Riffle
Affiliation:
Macromolecules and Interfaces Institute, Virginia Tech, Blacksburg, VA 24061, USA Department of Chemistry, Virginia Tech, Blacksburg, VA 24061, USA
Richey M Davis
Affiliation:
Macromolecules and Interfaces Institute, Virginia Tech, Blacksburg, VA 24061, USA Department of Chemical Engineering, Virginia Tech, Blacksburg, VA 24061, USA
Sarah M McDonald
Affiliation:
Virginia Tech Carilion Research Institute, Virginia Tech, Roanoke, VA 24016, USA
Deborah F Kelly
Affiliation:
Macromolecules and Interfaces Institute, Virginia Tech, Blacksburg, VA 24061, USA Virginia Tech Carilion Research Institute, Virginia Tech, Roanoke, VA 24016, USA
Corresponding
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Abstract

Understanding the fundamental properties of macromolecules has enhanced the development of emerging technologies used to improve biomedical research. Currently, there is a critical need for innovative platforms that can illuminate the function of biomedical reagents in a native environment. To address this need, we have developed an in situ approach to visualize the dynamic behavior of biomedically relevant macromolecules at the nanoscale. Newly designed silicon nitride devices containing integrated “microwells” were used to enclose active macromolecular specimens in liquid for transmission electron microscopy imaging purposes.We were able to successfully examine novel magnetic resonance imaging contrast reagents, micelle suspensions, liposome carrier vehicles, and transcribing viral assemblies. With each specimen tested, the integrated microwells adequately maintained macromolecules in discrete local environments while enabling thin liquid layers to be produced.

Type
In Situ Special Section
Copyright
© Microscopy Society of America 2014 

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Dukes et al. Supplementary Material

Dynamic behavior of magnetic nanoparticles in liquid. We examined microwells containing liquid and nanoparticles at low magnification then focused in on a selected microwell for a closer view. Small nanoparticle clusters (< 100 nm) can freely diffuse in solution while larger clusters (> 150 nm) remain relatively fixed. Smaller diffusing cluster tend to associate with the larger clusters over time, possibly by van der Waals attraction enhanced by the presence of iron oxide in the clusters.

Video 11 MB

Dukes et al. Supplementary Material

Micelles in solution lack long-range diffusion. An image sequence of micelles in solution show little to no diffusion in the X- and Y-directions although the micelles, which vary in size from 100 – 150 nm, may freely diffuse in the Z-direction and out of the focal plane.

Video 5 MB

Dukes et al. Supplementary Material

Structural details of DLPs in solution. A representative image of transcribing DLPs in solution was used to calculate 3D reconstructions (blue and yellow) of the active particles. Sections through the reconstructions reveal ordered density within the capsid cores at varying levels of transcriptional activity.

Video 5 MB

Dukes et al. Supplementary Material

Figures

PDF 9 MB

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