Mapping the dynamical organization of the cell nucleus through fluorescence correlation spectroscopy
The hierarchical organization of the cell nucleus into specialized open reservoirs and the nucleoplasm overcrowding impose restrictions to the mobility of biomolecules and their interactions with nuclear targets. These properties determine that many nuclear functions such as transcription, replicati...
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2018
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| Acceso en línea: | https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_10462023_v140-141_n_p10_Stortz http://hdl.handle.net/20.500.12110/paper_10462023_v140-141_n_p10_Stortz |
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paper:paper_10462023_v140-141_n_p10_Stortz2023-06-08T16:01:12Z Mapping the dynamical organization of the cell nucleus through fluorescence correlation spectroscopy Fluorescence correlation spectroscopy Fluorescence microscopy Glucocorticoid receptor Nucleus Simulations Transcription factor glucocorticoid receptor Article cell nucleus cytoplasm fluorescence correlation spectroscopy molecular dynamics molecular interaction nonhuman statistical analysis animal cell culture technique cell line cell nucleus devices fluorescence microscopy fluorescent antibody technique intravital microscopy laser laser scanning cytometry Mesocricetus metabolism procedures spectrofluorometry Animals Cell Culture Techniques Cell Line Cell Nucleus Fluorescent Antibody Technique Intravital Microscopy Laser Scanning Cytometry Lasers Mesocricetus Microscopy, Fluorescence Spectrometry, Fluorescence The hierarchical organization of the cell nucleus into specialized open reservoirs and the nucleoplasm overcrowding impose restrictions to the mobility of biomolecules and their interactions with nuclear targets. These properties determine that many nuclear functions such as transcription, replication, splicing or DNA repair are regulated by complex, dynamical processes that do not follow simple rules. Advanced fluorescence microscopy tools and, in particular, fluorescence correlation spectroscopy (FCS) provide complementary and exquisite information on the dynamics of fluorescent labeled molecules moving through the nuclear space and are helping us to comprehend the complexity of the nuclear structure. Here, we describe how FCS methods can be applied to reveal the dynamical organization of the nucleus in live cells. Specifically, we provide instructions for the preparation of cellular samples with fluorescent tagged proteins and detail how FCS can be easily instrumented in commercial confocal microscopes. In addition, we describe general rules to set the parameters for one and two-color experiments and the required controls for these experiments. Finally, we review the statistical analysis of the FCS data and summarize the use of numerical simulations as a complementary approach that helps us to understand the complex matrix of molecular interactions network within the nucleus. © 2017 Elsevier Inc. 2018 https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_10462023_v140-141_n_p10_Stortz http://hdl.handle.net/20.500.12110/paper_10462023_v140-141_n_p10_Stortz |
| institution |
Universidad de Buenos Aires |
| institution_str |
I-28 |
| repository_str |
R-134 |
| collection |
Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) |
| topic |
Fluorescence correlation spectroscopy Fluorescence microscopy Glucocorticoid receptor Nucleus Simulations Transcription factor glucocorticoid receptor Article cell nucleus cytoplasm fluorescence correlation spectroscopy molecular dynamics molecular interaction nonhuman statistical analysis animal cell culture technique cell line cell nucleus devices fluorescence microscopy fluorescent antibody technique intravital microscopy laser laser scanning cytometry Mesocricetus metabolism procedures spectrofluorometry Animals Cell Culture Techniques Cell Line Cell Nucleus Fluorescent Antibody Technique Intravital Microscopy Laser Scanning Cytometry Lasers Mesocricetus Microscopy, Fluorescence Spectrometry, Fluorescence |
| spellingShingle |
Fluorescence correlation spectroscopy Fluorescence microscopy Glucocorticoid receptor Nucleus Simulations Transcription factor glucocorticoid receptor Article cell nucleus cytoplasm fluorescence correlation spectroscopy molecular dynamics molecular interaction nonhuman statistical analysis animal cell culture technique cell line cell nucleus devices fluorescence microscopy fluorescent antibody technique intravital microscopy laser laser scanning cytometry Mesocricetus metabolism procedures spectrofluorometry Animals Cell Culture Techniques Cell Line Cell Nucleus Fluorescent Antibody Technique Intravital Microscopy Laser Scanning Cytometry Lasers Mesocricetus Microscopy, Fluorescence Spectrometry, Fluorescence Mapping the dynamical organization of the cell nucleus through fluorescence correlation spectroscopy |
| topic_facet |
Fluorescence correlation spectroscopy Fluorescence microscopy Glucocorticoid receptor Nucleus Simulations Transcription factor glucocorticoid receptor Article cell nucleus cytoplasm fluorescence correlation spectroscopy molecular dynamics molecular interaction nonhuman statistical analysis animal cell culture technique cell line cell nucleus devices fluorescence microscopy fluorescent antibody technique intravital microscopy laser laser scanning cytometry Mesocricetus metabolism procedures spectrofluorometry Animals Cell Culture Techniques Cell Line Cell Nucleus Fluorescent Antibody Technique Intravital Microscopy Laser Scanning Cytometry Lasers Mesocricetus Microscopy, Fluorescence Spectrometry, Fluorescence |
| description |
The hierarchical organization of the cell nucleus into specialized open reservoirs and the nucleoplasm overcrowding impose restrictions to the mobility of biomolecules and their interactions with nuclear targets. These properties determine that many nuclear functions such as transcription, replication, splicing or DNA repair are regulated by complex, dynamical processes that do not follow simple rules. Advanced fluorescence microscopy tools and, in particular, fluorescence correlation spectroscopy (FCS) provide complementary and exquisite information on the dynamics of fluorescent labeled molecules moving through the nuclear space and are helping us to comprehend the complexity of the nuclear structure. Here, we describe how FCS methods can be applied to reveal the dynamical organization of the nucleus in live cells. Specifically, we provide instructions for the preparation of cellular samples with fluorescent tagged proteins and detail how FCS can be easily instrumented in commercial confocal microscopes. In addition, we describe general rules to set the parameters for one and two-color experiments and the required controls for these experiments. Finally, we review the statistical analysis of the FCS data and summarize the use of numerical simulations as a complementary approach that helps us to understand the complex matrix of molecular interactions network within the nucleus. © 2017 Elsevier Inc. |
| title |
Mapping the dynamical organization of the cell nucleus through fluorescence correlation spectroscopy |
| title_short |
Mapping the dynamical organization of the cell nucleus through fluorescence correlation spectroscopy |
| title_full |
Mapping the dynamical organization of the cell nucleus through fluorescence correlation spectroscopy |
| title_fullStr |
Mapping the dynamical organization of the cell nucleus through fluorescence correlation spectroscopy |
| title_full_unstemmed |
Mapping the dynamical organization of the cell nucleus through fluorescence correlation spectroscopy |
| title_sort |
mapping the dynamical organization of the cell nucleus through fluorescence correlation spectroscopy |
| publishDate |
2018 |
| url |
https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_10462023_v140-141_n_p10_Stortz http://hdl.handle.net/20.500.12110/paper_10462023_v140-141_n_p10_Stortz |
| _version_ |
1768542798948073472 |