| Assay Method Information | |
| | n/a |
| Description: | Microsomal preparation: One lobe of fresh pig liver is obtained (e.g., at about the time of slaughter from a food-processing company) and immediately placed in ice cold 15 mM KH2PO4/250 mM sucrose (pH 7.4) and kept on ice during transportation. A 10 g sample of liver is minced and homogenized in 30 mL of homogenization buffer (15 mM KH2PO4/250 mM sucrose) using a Tekmar homoginizer or equivalent by pulsing 3 times with 20 second pulses. This procedure is repeated for a total of 8x10 g samples of pig liver. The remaining pig liver may be stored at -80 C. The homogenates from the 8 samples are pooled and centrifuged at 13,000xg for 20 minutes at 4 C. to remove crude debris. The supernatant is further centrifuged at 100,000xg for 70 minutes at 4° C. The microsomal pellets are re-suspended into 50 mL of 150 mM KH2PO4/1 mM DTT (pH 7.4) and 1 mL aliquots are stored at -80 C.100-150 ug of pig liver microsomal protein in 150 mM KH2PO4 is incubated at 37° C. |
| Affinity data for this assay | |
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