Acetate Incorporation into Lipid: Difference between revisions

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 # Turn radioactive tissue culture incubator on at 37C, make sure to turn on the CO2 as well.
 # Starve cells in '''Low Glucose Starvation Media''' for >3h.
-# Prepare '''Acetate Incorporation Media''' Make at least enough for one extra well, using 0.5 mL/well for a 12 well dish.  If monitoring lipogenesis in a line other than adipocytes, increase the hot glucose to 50 uCi per mL.
+# Prepare '''Acetate Incorporation Media''' Make at least enough for one extra well, using 50 uL/well for a 12 well dish.  If monitoring lipogenesis in a line other than adipocytes, increase the hot glucose to 50 uCi per mL.
 # Pretreat cells with inhibitors if required.
-# Change Media to '''Acetate Incorporation Media''' and add 100 nM insulin as required.
+# Add 100 nM insulin as required.
-# Place in the radioactive tissue culture incubator.
+# Add 50 uL of Hot Acetate Solution to each well.
+# Place in the radioactive tissue culture incubator for 60-120 minutes.
 # Save 3 x 5 uL of the '''Acetate Incorporation Media''' to count total radioactivity.
 # After 60 min  wash cells 3x with 1 mL of PBS.