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Comprehensive Method for Culturing Embryonic Dorsal Root Ganglion Neurons for Seahorse Extracellular Flux XF24 Analysis
Changes in mitochondrial dynamics and function contribute to progression of multiple neurodegenerative diseases including peripheral neuropathies. The Seahorse Extracellular Flux XF24 analyzer provides a comprehensive assessment of the relative state of glycolytic and aerobic metabolism in live cell...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Frontiers Media S.A.
2012
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3522103/ https://www.ncbi.nlm.nih.gov/pubmed/23248613 http://dx.doi.org/10.3389/fneur.2012.00175 |
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author | Lange, Miranda Zeng, Yan Knight, Andrew Windebank, Anthony Trushina, Eugenia |
author_facet | Lange, Miranda Zeng, Yan Knight, Andrew Windebank, Anthony Trushina, Eugenia |
author_sort | Lange, Miranda |
collection | PubMed |
description | Changes in mitochondrial dynamics and function contribute to progression of multiple neurodegenerative diseases including peripheral neuropathies. The Seahorse Extracellular Flux XF24 analyzer provides a comprehensive assessment of the relative state of glycolytic and aerobic metabolism in live cells making this method instrumental in assessing mitochondrial function. One of the most important steps in the analysis of mitochondrial respiration using the Seahorse XF24 analyzer is plating a uniform monolayer of firmly attached cells. However, culturing of primary dorsal root ganglion (DRG) neurons is associated with multiple challenges, including their propensity to form clumps and detach from the culture plate. This could significantly interfere with proper analysis and interpretation of data. We have tested multiple cell culture parameters including coating substrates, culture medium, XF24 microplate plastics, and plating techniques in order to optimize plating conditions. Here we describe a highly reproducible method to obtain neuron-enriched monolayers of securely attached dissociated primary embryonic (E15) rat DRG neurons suitable for analysis with the Seahorse XF24 platform. |
format | Online Article Text |
id | pubmed-3522103 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2012 |
publisher | Frontiers Media S.A. |
record_format | MEDLINE/PubMed |
spelling | pubmed-35221032012-12-17 Comprehensive Method for Culturing Embryonic Dorsal Root Ganglion Neurons for Seahorse Extracellular Flux XF24 Analysis Lange, Miranda Zeng, Yan Knight, Andrew Windebank, Anthony Trushina, Eugenia Front Neurol Neuroscience Changes in mitochondrial dynamics and function contribute to progression of multiple neurodegenerative diseases including peripheral neuropathies. The Seahorse Extracellular Flux XF24 analyzer provides a comprehensive assessment of the relative state of glycolytic and aerobic metabolism in live cells making this method instrumental in assessing mitochondrial function. One of the most important steps in the analysis of mitochondrial respiration using the Seahorse XF24 analyzer is plating a uniform monolayer of firmly attached cells. However, culturing of primary dorsal root ganglion (DRG) neurons is associated with multiple challenges, including their propensity to form clumps and detach from the culture plate. This could significantly interfere with proper analysis and interpretation of data. We have tested multiple cell culture parameters including coating substrates, culture medium, XF24 microplate plastics, and plating techniques in order to optimize plating conditions. Here we describe a highly reproducible method to obtain neuron-enriched monolayers of securely attached dissociated primary embryonic (E15) rat DRG neurons suitable for analysis with the Seahorse XF24 platform. Frontiers Media S.A. 2012-12-14 /pmc/articles/PMC3522103/ /pubmed/23248613 http://dx.doi.org/10.3389/fneur.2012.00175 Text en Copyright © 2012 Lange, Zeng, Knight, Windebank and Trushina. http://creativecommons.org/licenses/by/3.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in other forums, provided the original authors and source are credited and subject to any copyright notices concerning any third-party graphics etc. |
spellingShingle | Neuroscience Lange, Miranda Zeng, Yan Knight, Andrew Windebank, Anthony Trushina, Eugenia Comprehensive Method for Culturing Embryonic Dorsal Root Ganglion Neurons for Seahorse Extracellular Flux XF24 Analysis |
title | Comprehensive Method for Culturing Embryonic Dorsal Root Ganglion Neurons for Seahorse Extracellular Flux XF24 Analysis |
title_full | Comprehensive Method for Culturing Embryonic Dorsal Root Ganglion Neurons for Seahorse Extracellular Flux XF24 Analysis |
title_fullStr | Comprehensive Method for Culturing Embryonic Dorsal Root Ganglion Neurons for Seahorse Extracellular Flux XF24 Analysis |
title_full_unstemmed | Comprehensive Method for Culturing Embryonic Dorsal Root Ganglion Neurons for Seahorse Extracellular Flux XF24 Analysis |
title_short | Comprehensive Method for Culturing Embryonic Dorsal Root Ganglion Neurons for Seahorse Extracellular Flux XF24 Analysis |
title_sort | comprehensive method for culturing embryonic dorsal root ganglion neurons for seahorse extracellular flux xf24 analysis |
topic | Neuroscience |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3522103/ https://www.ncbi.nlm.nih.gov/pubmed/23248613 http://dx.doi.org/10.3389/fneur.2012.00175 |
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