Seahorse XF24, Руководство по обучению

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Basic Procedure
Contact Seahorse Technical Support: www.seahorsebio.com/techsupport
24
Oligomycin Optimization with the XF Cell Mito Stress Test
(Note: For this assay, seed cells at the optimal cell number that was determined previously.)
The XF Cell Mito Stress Test is run with four different concentrations of oligomycin to determine the
optimal oligomycin concentration to use in your XF assays. In a typical oligomycin optimization assay,
only three basal rate measurements followed by the oligomycin injection and three more rate
measurements need to be taken to determine the optimal concentration of oligomycin. However, for the
purposes of providing richer data for discussion, we will run the XF Cell Mito Stress Test and inject (A)
oligomycin (4 concentrations), (B) FCCP and (C) rotenone/antimycin A. In this experiment, we will use 0.5
µM
FCCP, although the optimal concentration of FCCP needs to be determined in a future assay.
Plate Layout:
Injections:
[Oligo mycin ] (µM)
0
0.5
1.0
2.0
All compounds will be made at 10x the final concentrations in the wells.
Port A: oligomycin
Row A : 0 µM final concentration in the well
Row B: 0.5 µM final concentration in the well
Row C: 1.0 µM final concentration in the well
Row D: 2.0 µM final concentration in the well
Port B: FCCP – 0.5
µM
final concentration in the well
Port C: rotenone/antimycin A – 0.5
µM
final concentration in the well