Novel Electron Tomographic Methods to Study the Morphology of Keratin Filament Networks
| Autor: | Paul Walther, Volker Schmidt, Katharina Höhn, Michaela Sailer, Michael Beil, Sebastian Lück |
|---|---|
| Rok vydání: | 2010 |
| Předmět: |
chemistry.chemical_classification
Electron Microscope Tomography Materials science Keratin Filament Macromolecular Substances Scanning electron microscope macromolecular substances law.invention Protein filament chemistry law Cell Line Tumor Keratin Microscopy Scanning transmission electron microscopy Biophysics Humans Keratins Electron microscope Intermediate filament Pancreas Instrumentation |
| Zdroj: | Microscopy and Microanalysis. 16:462-471 |
| ISSN: | 1435-8115 1431-9276 |
| DOI: | 10.1017/s1431927610093657 |
| Popis: | The three-dimensional (3D) keratin filament network of pancreatic carcinoma cells was investigated with different electron microscopical approaches. Semithin sections of high-pressure frozen and freeze substituted cells were analyzed with scanning transmission electron microscope (STEM) tomography. Preservation of subcellular structures was excellent, and keratin filaments could be observed; however, it was impossible to three-dimensionally track the individual filaments. To obtain a better signal-to-noise ratio in transmission mode, we observed ultrathin sections of high-pressure frozen and freeze substituted samples with low-voltage (30 kV) STEM. Contrast was improved compared to 300 kV, and individual filaments could be observed. The filament network of samples prepared by detergent extraction was imaged by high-resolution scanning electron microscopy (SEM) with very good signal-to-noise ratio using the secondary electron signal and the 3D structure could be elucidated by SEM tomography. In freeze-dried samples it was possible to discern between keratin filaments and actin filaments because the helical arrangement of actin subunits in the F-actin could be resolved. When comparing the network structures of the differently prepared samples, we found no obvious differences in filament length and branching, indicating that the intermediate filament network is less susceptible to preparation artifacts than the actin network. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
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