Fluorescence correlation spectroscopy experiments to quantify free diffusion coefficients in reaction-diffusion systems: The case of Ca2+ and its dyes
Many cell signaling pathways involve the diffusion of messengers that bind and unbind to and from intracellular components. Quantifying their net transport rate under different conditions then requires having separate estimates of their free diffusion coefficient and binding or unbinding rates. In t...
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| Acceso en línea: | https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_24700045_v95_n6_p_Sigaut http://hdl.handle.net/20.500.12110/paper_24700045_v95_n6_p_Sigaut |
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paper:paper_24700045_v95_n6_p_Sigaut2023-06-08T16:36:43Z Fluorescence correlation spectroscopy experiments to quantify free diffusion coefficients in reaction-diffusion systems: The case of Ca2+ and its dyes Sigaut, Lorena Ponce Dawson, Silvina Bins Calcium Cell signaling Dextran Diffusion in liquids Dynamic models Enzyme activity Fluorescence Fluorescence spectroscopy Physiological models Reaction rates Solutions Spectroscopic analysis Biophysical parameters Fluorescence Correlation Spectroscopy Intracellular components Net transport rate Reaction diffusion systems Relevant reactions Signaling pathways Xenopus laevis oocytes Diffusion Many cell signaling pathways involve the diffusion of messengers that bind and unbind to and from intracellular components. Quantifying their net transport rate under different conditions then requires having separate estimates of their free diffusion coefficient and binding or unbinding rates. In this paper, we show how performing sets of fluorescence correlation spectroscopy (FCS) experiments under different conditions, it is possible to quantify free diffusion coefficients and on and off rates of reaction-diffusion systems. We develop the theory and present a practical implementation for the case of the universal second messenger, calcium (Ca2+) and single-wavelength dyes that increase their fluorescence upon Ca2+ binding. We validate the approach with experiments performed in aqueous solutions containing Ca2+ and Fluo4 dextran (both in its high and low affinity versions). Performing FCS experiments with tetramethylrhodamine-dextran in Xenopus laevis oocytes, we infer the corresponding free diffusion coefficients in the cytosol of these cells. Our approach can be extended to other physiologically relevant reaction-diffusion systems to quantify biophysical parameters that determine the dynamics of various variables of interest. © 2017 American Physical Society. Fil:Sigaut, L. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. Fil:Ponce Dawson, S. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. 2017 https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_24700045_v95_n6_p_Sigaut http://hdl.handle.net/20.500.12110/paper_24700045_v95_n6_p_Sigaut |
| institution |
Universidad de Buenos Aires |
| institution_str |
I-28 |
| repository_str |
R-134 |
| collection |
Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) |
| topic |
Bins Calcium Cell signaling Dextran Diffusion in liquids Dynamic models Enzyme activity Fluorescence Fluorescence spectroscopy Physiological models Reaction rates Solutions Spectroscopic analysis Biophysical parameters Fluorescence Correlation Spectroscopy Intracellular components Net transport rate Reaction diffusion systems Relevant reactions Signaling pathways Xenopus laevis oocytes Diffusion |
| spellingShingle |
Bins Calcium Cell signaling Dextran Diffusion in liquids Dynamic models Enzyme activity Fluorescence Fluorescence spectroscopy Physiological models Reaction rates Solutions Spectroscopic analysis Biophysical parameters Fluorescence Correlation Spectroscopy Intracellular components Net transport rate Reaction diffusion systems Relevant reactions Signaling pathways Xenopus laevis oocytes Diffusion Sigaut, Lorena Ponce Dawson, Silvina Fluorescence correlation spectroscopy experiments to quantify free diffusion coefficients in reaction-diffusion systems: The case of Ca2+ and its dyes |
| topic_facet |
Bins Calcium Cell signaling Dextran Diffusion in liquids Dynamic models Enzyme activity Fluorescence Fluorescence spectroscopy Physiological models Reaction rates Solutions Spectroscopic analysis Biophysical parameters Fluorescence Correlation Spectroscopy Intracellular components Net transport rate Reaction diffusion systems Relevant reactions Signaling pathways Xenopus laevis oocytes Diffusion |
| description |
Many cell signaling pathways involve the diffusion of messengers that bind and unbind to and from intracellular components. Quantifying their net transport rate under different conditions then requires having separate estimates of their free diffusion coefficient and binding or unbinding rates. In this paper, we show how performing sets of fluorescence correlation spectroscopy (FCS) experiments under different conditions, it is possible to quantify free diffusion coefficients and on and off rates of reaction-diffusion systems. We develop the theory and present a practical implementation for the case of the universal second messenger, calcium (Ca2+) and single-wavelength dyes that increase their fluorescence upon Ca2+ binding. We validate the approach with experiments performed in aqueous solutions containing Ca2+ and Fluo4 dextran (both in its high and low affinity versions). Performing FCS experiments with tetramethylrhodamine-dextran in Xenopus laevis oocytes, we infer the corresponding free diffusion coefficients in the cytosol of these cells. Our approach can be extended to other physiologically relevant reaction-diffusion systems to quantify biophysical parameters that determine the dynamics of various variables of interest. © 2017 American Physical Society. |
| author |
Sigaut, Lorena Ponce Dawson, Silvina |
| author_facet |
Sigaut, Lorena Ponce Dawson, Silvina |
| author_sort |
Sigaut, Lorena |
| title |
Fluorescence correlation spectroscopy experiments to quantify free diffusion coefficients in reaction-diffusion systems: The case of Ca2+ and its dyes |
| title_short |
Fluorescence correlation spectroscopy experiments to quantify free diffusion coefficients in reaction-diffusion systems: The case of Ca2+ and its dyes |
| title_full |
Fluorescence correlation spectroscopy experiments to quantify free diffusion coefficients in reaction-diffusion systems: The case of Ca2+ and its dyes |
| title_fullStr |
Fluorescence correlation spectroscopy experiments to quantify free diffusion coefficients in reaction-diffusion systems: The case of Ca2+ and its dyes |
| title_full_unstemmed |
Fluorescence correlation spectroscopy experiments to quantify free diffusion coefficients in reaction-diffusion systems: The case of Ca2+ and its dyes |
| title_sort |
fluorescence correlation spectroscopy experiments to quantify free diffusion coefficients in reaction-diffusion systems: the case of ca2+ and its dyes |
| publishDate |
2017 |
| url |
https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_24700045_v95_n6_p_Sigaut http://hdl.handle.net/20.500.12110/paper_24700045_v95_n6_p_Sigaut |
| work_keys_str_mv |
AT sigautlorena fluorescencecorrelationspectroscopyexperimentstoquantifyfreediffusioncoefficientsinreactiondiffusionsystemsthecaseofca2anditsdyes AT poncedawsonsilvina fluorescencecorrelationspectroscopyexperimentstoquantifyfreediffusioncoefficientsinreactiondiffusionsystemsthecaseofca2anditsdyes |
| _version_ |
1768545440810139648 |