Using the Poseidon Select to Correlate Light and Electron Microscopy (CLEM) of Eukaryotic Cells in Liquid

Correlative Light and Electron Microscopy (CLEM) is an imaging technique that combines the functionality of light microscopy with the high resolution of electron microscopy. However, a major limitation of CLEM in biological applications is the need to dry or cryogenically freeze samples, which not only complicates experimental procedures, but is also likely to introduce artifacts.

The Poseidon System holder enables wet samples to be imaged with electron microscopy. Therefore, cellular receptors that are tagged with nanoparticles, such as quantum dots (QDs), can be imaged with nanometer resolution under physiologically relevant conditions.

Experiment

In this experiment, COS-7 fibroblast cells were directly cultured on Poseidon System E-chips™ sample supports. To label the epidermal growth factor (EGF) receptors, the cells were incubated for a period of 5 minutes in a 5 nM CdSe QDs solution coupled to EGF.

The cells were then washed, adhered with glutaraldehyde, and stored in phosphate buffered saline after labeling. The E-chips™ containing the cells were inverted and placed in a glass bottom culture dish to obtain fluorescence images.

A wide field fluorescence microscope fitted with an oil immersion objective was used to record all images. The E-chip™ was then placed in the Poseidon System holder with a liquid thickness of 5 µm and embedded into an FEI CM200 electron microscope. The sample was imaged in scanning TEM, and a 2 µL/min liquid flow rate was maintained during the entire imaging session.

Discussion

The electron microscopy images were correlated with their corresponding location in the light microscopy image. The E-chip™ window has a rectangular design that was used as a reference system for matching position coordinates between images.

The direct interference contrast image of a cell present on the E-chip™ window is shown in Figure A, and the equivalent quantum dot fluorescence is shown in Figure B. As shown in Figure C, the area indicated with an arrow was using scanning TEM.

The individual QDs can be seen as yellow-green spots spread all over the cell (false colorized to improve contrast). Cellular contours can be seen as regions of blue shading. By using the 25-75% edge width of line scans taken over 10 QDs, the spatial resolution was determined to be 3 nm.

Application

The study results show the efficiency of the Poseidon System for CLEM imaging of protein distribution on entire cells. The achieved resolution is adequate to distinguish different size, shape, and electron density among nanoparticles to facilitate complex imaging studies.

This information has been sourced, reviewed and adapted from materials provided by Protochips.

For more information on this source, please visit Protochips.

Citations

Please use one of the following formats to cite this article in your essay, paper or report:

  • APA

    Protochips. (2018, August 27). Using the Poseidon Select to Correlate Light and Electron Microscopy (CLEM) of Eukaryotic Cells in Liquid. AZoNano. Retrieved on June 24, 2019 from https://www.azonano.com/article.aspx?ArticleID=4334.

  • MLA

    Protochips. "Using the Poseidon Select to Correlate Light and Electron Microscopy (CLEM) of Eukaryotic Cells in Liquid". AZoNano. 24 June 2019. <https://www.azonano.com/article.aspx?ArticleID=4334>.

  • Chicago

    Protochips. "Using the Poseidon Select to Correlate Light and Electron Microscopy (CLEM) of Eukaryotic Cells in Liquid". AZoNano. https://www.azonano.com/article.aspx?ArticleID=4334. (accessed June 24, 2019).

  • Harvard

    Protochips. 2018. Using the Poseidon Select to Correlate Light and Electron Microscopy (CLEM) of Eukaryotic Cells in Liquid. AZoNano, viewed 24 June 2019, https://www.azonano.com/article.aspx?ArticleID=4334.

Ask A Question

Do you have a question you'd like to ask regarding this article?

Leave your feedback
Submit