Localisation of GFP and GFP tagged PKD2 in cultivated pancreatic cancer
                        cells using high-pressure freezing

Krisp, Holger; Wichert, Götz von; Seufferlein, Thomas; Walther, Paul

doi:10.1017/s1431927607080865

Cited by 1 publication

(1 citation statement)

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“…For fluorescence preserving preparation, the cells on the gridded sapphire discs were high pressure frozen, freeze substituted and finally embedded in LR-Gold resin using an adapted protocol introduced by Krisp et al (2007). After the polymerization, the coordinate system of the sapphire disc leaves an imprint on the surface of the polymerized resin block that serves as an accurate and stable reference system.…”

Section: Resultsmentioning

confidence: 99%

Preservation of protein fluorescence in embedded human dendritic cells for targeted 3D light and electron microscopy

Höhn

Fuchs

Fröber

et al. 2015

Journal of Microscopy

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SummaryIn this study, we present a correlative microscopy workflow to combine detailed 3D fluorescence light microscopy data with ultrastructural information gained by 3D focused ion beam assisted scanning electron microscopy. The workflow is based on an optimized high pressure freezing/freeze substitution protocol that preserves good ultrastructural detail along with retaining the fluorescence signal in the resin embedded specimens. Consequently, cellular structures of interest can readily be identified and imaged by state of the art 3D confocal fluorescence microscopy and are precisely referenced with respect to an imprinted coordinate system on the surface of the resin block. This allows precise guidance of the focused ion beam assisted scanning electron microscopy and limits the volume to be imaged to the structure of interest. This, in turn, minimizes the total acquisition time necessary to conduct the time consuming ultrastructural scanning electron microscope imaging while eliminating the risk to miss parts of the target structure. We illustrate the value of this workflow for targeting virus compartments, which are formed in HIV‐pulsed mature human dendritic cells.

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Section: Resultsmentioning

confidence: 99%

Preservation of protein fluorescence in embedded human dendritic cells for targeted 3D light and electron microscopy

Höhn

Fuchs

Fröber

et al. 2015

Journal of Microscopy

View full text Add to dashboard Cite

show abstract

Localisation of GFP and GFP tagged PKD2 in cultivated pancreatic cancer cells using high-pressure freezing

Abstract: Extended abstract of a paper presented at MC 2007, 33rd DGE Conference in Saarbrücken, Germany, September 2 – September 7, 2007

Cited by 1 publication

References 3 publications

Preservation of protein fluorescence in embedded human dendritic cells for targeted 3D light and electron microscopy

Preservation of protein fluorescence in embedded human dendritic cells for targeted 3D light and electron microscopy

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