FLIPPER, a combinatorial probe for correlated live imaging and electron microscopy, allows identification and quantitative analysis of various cells and organelles

Cell and Tissue Research
Jeroen KuipersBen N G Giepmans

Abstract

Ultrastructural examination of cells and tissues by electron microscopy (EM) yields detailed information on subcellular structures. However, EM is typically restricted to small fields of view at high magnification; this makes quantifying events in multiple large-area sample sections extremely difficult. Even when combining light microscopy (LM) with EM (correlated LM and EM: CLEM) to find areas of interest, the labeling of molecules is still a challenge. We present a new genetically encoded probe for CLEM, named "FLIPPER", which facilitates quantitative analysis of ultrastructural features in cells. FLIPPER consists of a fluorescent protein (cyan, green, orange, or red) for LM visualization, fused to a peroxidase allowing visualization of targets at the EM level. The use of FLIPPER is straightforward and because the module is completely genetically encoded, cells can be optimally prepared for EM examination. We use FLIPPER to quantify cellular morphology at the EM level in cells expressing a normal and disease-causing point-mutant cell-surface protein called EpCAM (epithelial cell adhesion molecule). The mutant protein is retained in the endoplasmic reticulum (ER) and could therefore alter ER function and morphology. To reveal ...Continue Reading

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Citations

May 29, 2015·Nature Methods·Pascal de BoerBen N G Giepmans
Jan 27, 2015·Cell and Tissue Research·Tania Q VuDiane S Lidke
Mar 17, 2015·Cell and Tissue Research·Christopher S von Bartheld, Fred S Wouters
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Mar 20, 2019·Journal of Cell Science·Ranjan SenguptaSeema Mattoo
Jan 13, 2018·Histochemistry and Cell Biology·Marit A de BeerBen N G Giepmans
Dec 7, 2018·Proceedings of the National Academy of Sciences of the United States of America·Julia K DohKimberly E Beatty
Nov 27, 2020·Frontiers in Cellular Neuroscience·Marit A de Beer, Ben N G Giepmans
Mar 9, 2021·Current Opinion in Cell Biology·Kem A Sochacki, Justin W Taraska

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Methods Mentioned

BETA
light microscopy
fluorescence microscopy
transfection
light
electron microscopy
confocal microscopy

Software Mentioned

SOG
mini
CLEM
Adobe Photoshop
ImageJ Fiji
Image J Fiji

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