Triglyceride Assay from Cells and Tissues: Difference between revisions
updated volumes and added a new section describing tissue specific volumes to use |
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==Materials== | ==Materials== | ||
* '''Homogenization Buffer''' (50 mM Tris, 5 mM EDTA, 30 mM Mannitol, PI inhibitor) | * '''Homogenization Buffer''' (50 mM Tris, 5 mM EDTA, 30 mM Mannitol, PI inhibitor, can be made in bulk without the PI, PI added fresh) | ||
* 10M KOH | * 10M KOH | ||
* '''Chloroform/Methanol Mixture''' (2:1) | * '''Chloroform/Methanol Mixture''' (2:1) | ||
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==Protocol== | ==Protocol== | ||
# Weigh out 200-500mg tissue (record weight for normalization). You can use | # Weigh out 200-500mg tissue (record weight for normalization). You can use as low as 30 mg tissue if necessary by reducing the lysis volume (must be at least 500 uL) or increasing the amount of the lower (chloroform) layer removed up to 500 ul(normally 180 uL). | ||
# Homogenize with dounce homogenizer in 2 mL '''Homogenization Buffer'''. | # Homogenize with dounce homogenizer in 2 mL '''Homogenization Buffer'''. | ||
# Remove 200 uL to a tube containing 5 uL KOH | # Remove 200 uL to a tube containing 5 uL KOH | ||