| AVS 58th Annual International Symposium and Exhibition | |
| In Situ Spectroscopy and Microscopy Focus Topic | Tuesday Sessions |
| Session IS+AS+SS-TuM |
| Session: | In Situ Studies of Organic and Soft Materials and Liquid-Solid Interfaces |
| Presenter: | Niels De Jonge, Vanderbilt University School of Medicine |
| Authors: | N. De Jonge, Vanderbilt University School of Medicine D.B. Peckys, Vanderbilt University School of Medicine |
| Correspondent: | Click to Email |
We have recently introduced a novel electron microscopy technique for the imaging of whole cells in aqueous media using scanning transmission electron microscopy (STEM) [1, 2]. Eukaryotic cells in liquid were placed in a microfluidic chamber with a thickness of 5 - 10 μm contained between two ultra-thin electron-transparent windows. On account of the atomic number (Z) contrast of the STEM, nanoparticles of a high-Z material (e.g., gold) were detected within the background signal produced by a micrometers-thick layer of a low-Z liquid (e.g. water, or cellular material). Nanoparticles specifically attached to proteins can be used to study protein distributions in whole cells in liquid, similar as proteins tagged with fluorescent labels can be used to study protein distributions in cells with fluorescence microscopy.
COS7 fibroblast cells were labeled with gold nanoparticles conjugated with epidermal growth factor (EGF). Intact fixed cells in liquid were imaged with STEM with a spatial resolution of 4 nm and a pixel dwell time of 20 microseconds [1]. In test experiments we demonstrated a maximal spatial resolution of 1.5 nm on gold nanoparticles placed above a water layer of a thickness of 3 micrometer, consistent with theoretical predictions, and with Monte Carlo simulations of the STEM imaging [3]. The use of quantum dots (QDs), which are fluorescent nanoparticles, allowed STEM images to be correlated with fluorescence images [4]. Eukaryotic cells were grown directly on microchips for the microfluidic chamber, fixed, and imaged with fluorescence microscopy. The intact cells were then imaged in liquid with STEM. The STEM images showed individual QDs, and their locations were correlated with the cellular regions, as imaged with fluorescence microscopy. We have also demonstrated the imaging of nanoparticle uptake in live cells [5], and the ultrastructure of pristine yeast cells was studied [6]. Liquid STEM presents an innovative approach for the imaging of whole cells, with significantly improved spatial resolution and imaging speed over existing methods.
URL: http://www.mc.vanderbilt.edu/labs/dejongelab/
References
[1] de Jonge, N., Peckys, D.B., Kremers, G.J. & Piston, D.W., Proc. Natl. Acad. Sci. 106, 2159-2164, 2009.
[2] Peckys, D.B., Veith, G.M., Joy, D.C. & de Jonge, N., PLoS One 4, e8214-1-7, 2009.
[3] Dukes, M.J., Peckys, D.B. & de Jonge, N., ACS Nano 4, 4110-4116, 2010.
[4] de Jonge, N., Poirier-Demers, N., Demers, H., Peckys, D.B. & Drouin, D., Ultramicroscopy 110, 1114-1119, 2010.
[5] Peckys, D.B. & N. de Jonge, Nano Lett. 11, 1733-1738, 2011.
[6] Peckys, D.B., Mazur, P., Gould, K.L. & de Jonge, N., Biophys. J., in press, 2011.