Challenges of Using Expansion Microscopy for Super‐resolved Imaging of Cellular Organelles
| Autor: | Christoffer Lagerholm, Christian Eggeling, Silvia Galiani, Dominic Waithe, Wolfgang Schliebs, Katharina Reglinski, Ralf Erdmann, Maximilian Büttner |
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| Rok vydání: | 2020 |
| Předmět: |
cell organelles
Matrix (biology) 010402 general chemistry 01 natural sciences Biochemistry law.invention Very Important Paper STED microscopy Confocal microscopy law Microscopy Organelle Peroxisomes Humans bioorganic chemistry Molecular Biology Polyacrylamide gel electrophoresis Cellular compartment Cell Nucleus Microscopy Confocal Full Paper 010405 organic chemistry Chemistry Cell Membrane Organic Chemistry Full Papers Peroxisome Mitochondria Molecular Imaging 0104 chemical sciences HEK293 Cells Microscopy Fluorescence peptides Biophysics Molecular Medicine expansion microscopy |
| Zdroj: | Chembiochem |
| ISSN: | 1439-7633 1439-4227 |
| Popis: | Expansion microscopy (ExM) has been successfully used to improve the spatial resolution when imaging tissues by optical microscopy. In ExM, proteins of a fixed sample are crosslinked to a swellable acrylamide gel, which expands when incubated in water. Therefore, ExM allows enlarged subcellular structures to be resolved that would otherwise be hidden to standard confocal microscopy. Herein, we aim to validate ExM for the study of peroxisomes, mitochondria, nuclei and the plasma membrane. Upon comparison of the expansion factors of these cellular compartments in HEK293 cells within the same gel, we found significant differences, of a factor of above 2, in expansion factors. For peroxisomes, the expansion factor differed even between peroxisomal membrane and matrix marker; this underlines the need for a thorough validation of expansion factors of this powerful technique. We further give an overview of possible quantification methods for the determination of expansion factors of intracellular organelles, and we highlight some potentials and challenges. In Expansion Microscopy (ExM) subcellular structures are imaged in isotropically expanded fixed samples, consequently allowing to resolve enlarged subcellular structures, otherwise hidden to standard microscopy. Upon comparison of the expansion factors of different cellular compartments in cells within the same gel, we found significant differences in expansion factors of a factor of above 2. |
| Databáze: | OpenAIRE |
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