INS-1, MIN-6, HeLa and EA.hy926 cells, treated or not with siRNA against presenilin-1 or GSK3β inhibitor CHIR99021, were plated in XF96 polystyrene cell culture microplates (Seahorse Bioscience) at a density of 60.000 cells/well for HeLa and EA.hy926, 120.000 cells/well for MIN-6 and 140.000 cells/well for INS-1 cells. After overnight incubation, mitochondrial respiration was performed using an XF96 extracellular flux analyzer (Seahorse Bioscience®) as previously described [21 (link)]. Oxygen consumption was normalized to protein content (pmol O2/min × μg protein).
Protocol detail
Mitochondrial Respiration in Cell Lines
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Cell culture
Cells
Chir99021
Gsk3β
Hela
Mitochondrial
Oxygen consumption
Polystyrene
Presenilin 1
Protein
Respiration
Seahorse
Sirna
Corresponding Organization :
Other organizations : Medical University of Graz, University of Amsterdam, Academic Medical Center, University of Montana
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Variable analysis
independent variables
- SiRNA against presenilin-1
- GSK3β inhibitor CHIR99021
- Mitochondrial respiration (Oxygen consumption normalized to protein content (pmol O2/min × μg protein))
- Cell types: INS-1, MIN-6, HeLa and EA.hy926 cells
- Cell plating densities: 60,000 cells/well for HeLa and EA.hy926, 120,000 cells/well for MIN-6, and 140,000 cells/well for INS-1 cells
- Overnight incubation
- Mitochondrial respiration measurement using an XF96 extracellular flux analyzer
- Untreated (no siRNA against presenilin-1, no GSK3β inhibitor CHIR99021) cells for each cell type
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