View source for Measurement of Glycolysis and Respiration Using Seahorse

==Materials==
*Non Buffered DMEM - Sigma cat#D5648  Dissolve one bottle (13.4g) in a litre of water.  Adjust pH to 7-7.2 and filter into a sterile bottle.
*FCCP - Prepare as 300 uM stock in DMSO.  Need about 20 uL per plate
*Oligomycin - Prepare as a 1 mM Stock in DMSO.  Need about 20 uL per plate
*XF24 plate


==Protocol==
#Email Sydney Bridges to arrange a time to do the experiment.  His email is sbridges@med.umich.edu
#Trypsinise cells and resuspend in ~10 mL (for a 10 cm dish).
#Count cells using hemocytometer or cell counter.
#Adjust cell concentration to be 250 000 cells/mL (50K cells/200 uL) using normal growth media
#Add 200 uL of cell suspension per well of a XF24 plate.
#Let cells adhere for > 30min and then add 800 uL of buffer
#Incubate cells overnight.
#Prior to experiment, aspirate 950 uL media then add 1 mL non-buffered DMEM.
#Remove 1 mL media (leaving 50 uL)
#Add 625 uL of non-buffered DMEM for a final volume of 675 uL.
#Bring plate over to Brehm Room 6470 and place in non-CO2 incubator.