Comparing super-resolution microscopy techniques to analyze chromosomes
| Autor: | Eva Hřibová, Alžběta Němečková, Veit Schubert, Ivona Kubalová, Klaus Weisshart |
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| Jazyk: | angličtina |
| Rok vydání: | 2021 |
| Předmět: |
0106 biological sciences
0301 basic medicine Indoles structured illumination microscopy nanoscopy 01 natural sciences law.invention lcsh:Chemistry metaphase chromosome stimulated emission depletion microscopy law Microscopy photoactivated localization microscopy lcsh:QH301-705.5 Spectroscopy chromatin Hordeum vulgare wide-field microscopy topoisomerase II deconvolution microscopy Microscopy Confocal Super-resolution microscopy Resolution (electron density) STED microscopy General Medicine Single Molecule Imaging Computer Science Applications Materials science Catalysis Chromosomes Plant Article Inorganic Chemistry 03 medical and health sciences Confocal microscopy Photoactivated localization microscopy Physical and Theoretical Chemistry Molecular Biology Nanoscopic scale Metaphase Fluorescent Dyes Organic Chemistry Reproducibility of Results Hordeum 030104 developmental biology DNA Topoisomerases Type II Microscopy Fluorescence lcsh:Biology (General) lcsh:QD1-999 Biophysics 010606 plant biology & botany |
| Zdroj: | International journal of molecular sciences, 22(4):1903 International Journal of Molecular Sciences International Journal of Molecular Sciences, Vol 22, Iss 1903, p 1903 (2021) Volume 22 Issue 4 |
| Popis: | The importance of fluorescence light microscopy for understanding cellular and sub-cellular structures and functions is undeniable. However, the resolution is limited by light diffraction (~200–250 nm laterally, ~500–700 nm axially). Meanwhile, super-resolution microscopy, such as structured illumination microscopy (SIM), is being applied more and more to overcome this restriction. Instead, super-resolution by stimulated emission depletion (STED) microscopy achieving a resolution of ~50 nm laterally and ~130 nm axially has not yet frequently been applied in plant cell research due to the required specific sample preparation and stable dye staining. Single-molecule localization microscopy (SMLM) including photoactivated localization microscopy (PALM) has not yet been widely used, although this nanoscopic technique allows even the detection of single molecules. In this study, we compared protein imaging within metaphase chromosomes of barley via conventional wide-field and confocal microscopy, and the sub-diffraction methods SIM, STED, and SMLM. The chromosomes were labeled by DAPI (4′,6-diamidino-2-phenylindol), a DNA-specific dye, and with antibodies against topoisomerase IIα (Topo II), a protein important for correct chromatin condensation. Compared to the diffraction-limited methods, the combination of the three different super-resolution imaging techniques delivered tremendous additional insights into the plant chromosome architecture through the achieved increased resolution. |
| Databáze: | OpenAIRE |
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