High-content fluorescence imaging with the metabolic flux assay reveals insights into mitochondrial properties and functions
Autor: | Vinee Purohit, Sofia D. Merajver, David B. Lombard, Laura E. Goo, Ilya Kovalenko, Joel A. Yates, Samuel A. Kerk, Andrew C. Little, Hanna Sungok Hong, Costas A. Lyssiotis |
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Rok vydání: | 2020 |
Předmět: |
Fluorescence-lifetime imaging microscopy
Bioenergetics Medicine (miscellaneous) Breast Neoplasms Biochemistry General Biochemistry Genetics and Molecular Biology Article Cell Line Membrane Potentials 03 medical and health sciences 0302 clinical medicine Metabolic flux analysis Humans Glycolysis Fragmentation (cell biology) lcsh:QH301-705.5 Pancreas Sensors and probes 030304 developmental biology 0303 health sciences Chemistry Optical Imaging Metabolic Flux Analysis Mitochondria Metabolic pathway lcsh:Biology (General) Metabolic pathways 030220 oncology & carcinogenesis Biophysics General Agricultural and Biological Sciences Reactive Oxygen Species Flux (metabolism) Function (biology) |
Zdroj: | Communications Biology Communications Biology, Vol 3, Iss 1, Pp 1-10 (2020) |
ISSN: | 2399-3642 |
Popis: | Metabolic flux technology with the Seahorse bioanalyzer has emerged as a standard technique in cellular metabolism studies, allowing for simultaneous kinetic measurements of respiration and glycolysis. Methods to extend the utility and versatility of the metabolic flux assay would undoubtedly have immediate and wide-reaching impacts. Herein, we describe a platform that couples the metabolic flux assay with high-content fluorescence imaging to simultaneously provide means for normalization of respiration data with cell number; analyze cell cycle distribution; and quantify mitochondrial content, fragmentation state, membrane potential, and mitochondrial reactive oxygen species. Integration of fluorescent dyes directly into the metabolic flux assay generates a more complete data set of mitochondrial features in a single assay. Moreover, application of this integrated strategy revealed insights into mitochondrial function following PGC1a and PRC1 inhibition in pancreatic cancer and demonstrated how the Rho-GTPases impact mitochondrial dynamics in breast cancer. Little, Kovalenko et al. couple the metabolic flux assay from the Seahorse bioanalyzer with high-content fluorescence imaging to study the coordinated biology of mitochondrial bioenergetics and properties. They demonstrate this integration generates a more comprehensive data set, allowing for further insight into mitochondrial function and dynamics. |
Databáze: | OpenAIRE |
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