Triglyceride Assay from Cells and Tissues

Materials

• Homogenization Buffer (50 mM Tris pH 8, 5 mM EDTA, 30 mM Mannitol, PI inhibitor, can be made in bulk without the PI, PI added fresh)
• 10M KOH
• Chloroform/Methanol Mixture (2:1)
• Butanol Mixture: 3 mL butanol, 1.66 mL Triton-X114, 0.33 mL Methanol
• Sigma Triglyceride Assay Kit (Cat TR0100)

Protocol

1. Use 5 female or 8 Male flies for Assay
2. Homogenize flies (3 min @ 40Hz) in 1mL of .05% Tween (from Tween-20 stock)
3. Heat samples in 70C waterbath for 5 minutes
4. Spin samples @ 5000G for 1 minute
5. Transfer 500ul of supernatent in new microfuge tube
6. Spin @ 14000G for 3 minutes
7. Add 50ul of sample to 200ul of TG solution in a 96 well plate
8. Incubate plate @ 37C for 5 mins
9. Measure 540nm absorbance