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Measurement and Analysis of Extracellular Acid Production to Determine Glycolytic Rate
Extracellular measurement of oxygen consumption and acid production is a simple and powerful way to monitor rates of respiration and glycolysis(1). Both mitochondrial (respiration) and non-mitochondrial (other redox) reactions consume oxygen, but these reactions can be easily distinguished by chemic...
Autores principales: | , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MyJove Corporation
2015
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4692795/ https://www.ncbi.nlm.nih.gov/pubmed/26709455 http://dx.doi.org/10.3791/53464 |
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author | Mookerjee, Shona A. Brand, Martin D. |
author_facet | Mookerjee, Shona A. Brand, Martin D. |
author_sort | Mookerjee, Shona A. |
collection | PubMed |
description | Extracellular measurement of oxygen consumption and acid production is a simple and powerful way to monitor rates of respiration and glycolysis(1). Both mitochondrial (respiration) and non-mitochondrial (other redox) reactions consume oxygen, but these reactions can be easily distinguished by chemical inhibition of mitochondrial respiration. However, while mitochondrial oxygen consumption is an unambiguous and direct measurement of respiration rate(2), the same is not true for extracellular acid production and its relationship to glycolytic rate (3-6). Extracellular acid produced by cells is derived from both lactate, produced by anaerobic glycolysis, and CO(2), produced in the citric acid cycle during respiration. For glycolysis, the conversion of glucose to lactate(-) + H(+) and the export of products into the assay medium is the source of glycolytic acidification. For respiration, the export of CO(2), hydration to H(2)CO(3) and dissociation to HCO(3)(-) + H(+) is the source of respiratory acidification. The proportions of glycolytic and respiratory acidification depend on the experimental conditions, including cell type and substrate(s) provided, and can range from nearly 100% glycolytic acidification to nearly 100% respiratory acidification (6). Here, we demonstrate the data collection and calculation methods needed to determine respiratory and glycolytic contributions to total extracellular acidification by whole cells in culture using C2C12 myoblast cells as a model. |
format | Online Article Text |
id | pubmed-4692795 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2015 |
publisher | MyJove Corporation |
record_format | MEDLINE/PubMed |
spelling | pubmed-46927952016-01-07 Measurement and Analysis of Extracellular Acid Production to Determine Glycolytic Rate Mookerjee, Shona A. Brand, Martin D. J Vis Exp Biochemistry Extracellular measurement of oxygen consumption and acid production is a simple and powerful way to monitor rates of respiration and glycolysis(1). Both mitochondrial (respiration) and non-mitochondrial (other redox) reactions consume oxygen, but these reactions can be easily distinguished by chemical inhibition of mitochondrial respiration. However, while mitochondrial oxygen consumption is an unambiguous and direct measurement of respiration rate(2), the same is not true for extracellular acid production and its relationship to glycolytic rate (3-6). Extracellular acid produced by cells is derived from both lactate, produced by anaerobic glycolysis, and CO(2), produced in the citric acid cycle during respiration. For glycolysis, the conversion of glucose to lactate(-) + H(+) and the export of products into the assay medium is the source of glycolytic acidification. For respiration, the export of CO(2), hydration to H(2)CO(3) and dissociation to HCO(3)(-) + H(+) is the source of respiratory acidification. The proportions of glycolytic and respiratory acidification depend on the experimental conditions, including cell type and substrate(s) provided, and can range from nearly 100% glycolytic acidification to nearly 100% respiratory acidification (6). Here, we demonstrate the data collection and calculation methods needed to determine respiratory and glycolytic contributions to total extracellular acidification by whole cells in culture using C2C12 myoblast cells as a model. MyJove Corporation 2015-12-12 /pmc/articles/PMC4692795/ /pubmed/26709455 http://dx.doi.org/10.3791/53464 Text en Copyright © 2015, Journal of Visualized Experiments http://creativecommons.org/licenses/by-nc-nd/3.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs 3.0 Unported License. To view a copy of this license, visithttp://creativecommons.org/licenses/by-nc-nd/3.0/ |
spellingShingle | Biochemistry Mookerjee, Shona A. Brand, Martin D. Measurement and Analysis of Extracellular Acid Production to Determine Glycolytic Rate |
title | Measurement and Analysis of Extracellular Acid Production to Determine Glycolytic Rate |
title_full | Measurement and Analysis of Extracellular Acid Production to Determine Glycolytic Rate |
title_fullStr | Measurement and Analysis of Extracellular Acid Production to Determine Glycolytic Rate |
title_full_unstemmed | Measurement and Analysis of Extracellular Acid Production to Determine Glycolytic Rate |
title_short | Measurement and Analysis of Extracellular Acid Production to Determine Glycolytic Rate |
title_sort | measurement and analysis of extracellular acid production to determine glycolytic rate |
topic | Biochemistry |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4692795/ https://www.ncbi.nlm.nih.gov/pubmed/26709455 http://dx.doi.org/10.3791/53464 |
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