Steindel, M., Orsine de Almeida, I., Strawbridge, S. orcid.org/0000-0003-0273-0835 et al. (4 more authors) (2022) Studying the dynamics of chromatin-binding proteins in mammalian cells using single-molecule localization microscopy. In: Chromosome Architecture: Methods and Protocols. Methods in Molecular Biology, MIMB 2476 . Humana New York, NY , pp. 209-247. ISBN 9781071622209
Abstract
Single-molecule localization microscopy (SMLM) allows the super-resolved imaging of proteins within mammalian nuclei at spatial resolutions comparable to that of a nucleosome itself (~20 nm). The technique is therefore well suited to the study of chromatin structure. Fixed-cell SMLM has already allowed temporal "snapshots" of how proteins are arranged on chromatin within mammalian nuclei. In this chapter, we focus on how recent developments, for example in selective plane illumination, 3D SMLM, and protein labeling, have led to a range of live-cell SMLM studies. We describe how to carry out single-particle tracking (SPT) of single proteins and, by analyzing their diffusion parameters, how to determine whether proteins interact with chromatin, diffuse freely, or do both. We can study the numbers of proteins that interact with chromatin and also determine their residence time on chromatin. We can determine whether these proteins form functional clusters within the nucleus as well as whether they form specific nuclear structures.
Metadata
Item Type: | Book Section |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © 2022 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature. |
Keywords: | Anomalous exponent; Chromatin; Diffusion coefficient; Fluorescence imaging; Jump distance; Mean squared displacement; PALM; Residence time; SPIM; SPT; STORM; Super-resolution microscopy; Animals; Carrier Proteins; Chromatin; Chromosomes; Mammals; Microscopy; Single Molecule Imaging |
Dates: |
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Institution: | The University of Sheffield |
Academic Units: | The University of Sheffield > Faculty of Science (Sheffield) > School of Biosciences (Sheffield) |
Depositing User: | Symplectic Sheffield |
Date Deposited: | 21 Feb 2025 14:23 |
Last Modified: | 21 Feb 2025 14:23 |
Status: | Published |
Publisher: | Humana New York, NY |
Series Name: | Methods in Molecular Biology |
Refereed: | Yes |
Identification Number: | 10.1007/978-1-0716-2221-6_16 |
Related URLs: | |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:223519 |