Agilent Technologies
Seahorse XFe96 FluxPaks
102416-100
The research aim of this current study is to test how lipid regulates cell metabolism. Specifically, in this experiment, we aim to study how fatty acids regulate CD8 T cell mitochondrial oxidative phosphorylation. Briefly, T cells were seeded in the plate and loaded to the seahorse analyzer. Mitochondrial drugs were injected in the following order: oligomycin, FCCP, Etomoxir (an inhibitor of fatty acid transportation to mitochondria and Rotenone). Oxygen consumption rate (OCR) was monitored. Seahorse Bioscience (with Agilent Technologies) offers a very convenient tool to address this question, and that is why I choose this product for my experiment.
Seahorse extracelluar flux assay
Mouse T cells
1. T cells were seeded in the seahorse plate (96 well) in seahorse medium;2. Mitochondrial drugs (oligomycin, FCCP, Etomoxir and Rotenone) were pre-injected to the four portals on top of the plates. The equilibration takes about 25 minutes. 3. Replace the holder plate with the seahorse plate containing T cells, which will be automatically loaded to the seahorse machine.4. Oxygen consumption rate (OCR) values will display on the monitor screen. The total assay time depends on the program.
Be careful of the barcode of the plate.
As shown in the attached figure, oligomycin suppresses oxygen consumption, which is reversed and even further increased upon treatment with uncoupler FCCP. Etomoxir, which is an inhibitor of fatty acid transportation to mitochonria, substantially reduces the OCR, suggesting that T cells undergo fatty acid oxidation.
Very easy to use.
None
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Easy to use.
A little expensive.
In general, the product fulfill my experimental purpose. Although it is a little expensive, it is easy to use and the experimental result is of some value for my project.