Assessing isoproterenol-stimulated whole-body lipolysis in vivo
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Assessing Isoproterenol-Stimulated Whole-Body Lipolysis in vivo
- Note: Mice do not need to be in a fasted state prior to this test.
- Briefly anesthetize mice with isofluorane and collect blood via retro orbital bleed. Allow blood to clot over ice.
- Inject 10 mg/kg isoproterenol (prepared fresh in sterile PBS) via intraperitoneal injection.
- Wait 15 minutes.
- Briefly anesthetize mice with isoflurane and collect blood via retro orbital bleed. Allow blood to clot over ice.
- Centrifuge blood samples and collect serum.
- Assay serum for NEFA's and Glycerol/Triglycerides using Wako and Sigma assay kits, respectively.
NEFA determination from serum
- To wells of a clear 96 well plate, add 2.5 uL of water (as blank), appropriate standard (the kit comes with stock of 1 mEq/L. Dilute the stock in ddH20 to give 2.5 and 5 mEq/L standards. For the high standard, add 5 uL of the 1 mEq/L stock. Due to the extra volume, the concentration of this last standard is actually 1.97 mEq/L).
- To remaining wells, add 2.5 uL of serum samples.
- To each well, add 100 uL of reaction buffer A
