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MB Sample ID: SA217403
Local Sample ID: | Tbx3_KO_IF_1 |
Subject ID: | SU002349 |
Subject Type: | Mammal |
Subject Species: | Mus musculus |
Taxonomy ID: | 10090 |
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Sample Preparation:
Sampleprep ID: | SP002355 |
Sampleprep Summary: | Liver specimens were harvested and immediately flash frozen in LN2 then stored at -80°C. While kept on dry ice a 20 mg sample was removed from each liver specimen, massed using an analytical balance, and placed in a 2 mL round bottom polypropylene tube containing 4-6, 2.3 mm stainless steel beads. 500 µL of -20°C extraction solution (methanol: acetonitrile: water, 2:2:1) containing stable isotope labeled metabolite standards was added to each sample tube. Ratio of 20 mg to 500 µL was retained when masses were not exactly 20 mg. All samples were homogenized at an amplitude of 20 Hz for 15 minutes and stored at -20°C for one hour to maximize protein precipitation. Samples were then vortexed for 20 seconds and centrifuged at 4°C for 5 minutes, speed 14,000 rcf. 120 µL of supernatant was removed from each tube and filtered using 0.2 µm polyvinylidene fluoride filter (Agilent Technologies P/N: 203980-100) and collected via 6,000 rcf centrifuge for 4 minutes. An additional 50uL was removed from each sample and combined into 5 pooled samples analyzed at equal intervals throughout the analysis to ensure stable signal. Extracts, pools, and procedural blanks were sealed and stored at 4°C until prompt analysis. |