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MB Sample ID: SA272232
Local Sample ID: | RF164 |
Subject ID: | SU002812 |
Subject Type: | Mammal |
Subject Species: | Mus musculus |
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Sample Preparation:
Sampleprep ID: | SP002818 |
Sampleprep Summary: | CSF was collected from adult (3 months old) wild-type CD1 mice. Samples were placed on wet ice, then spun 1000xg for 10 minutes at 4C. The supernatant was collected. Per condition, 5-10µL of fresh, cleared CSF or 5µl serum was extracted in 4:6:3 chlorophorm:methanol:water mixture supplemented with isotopically labeled T3 and T4 (Cambridge Isotope Laboratories, CLM-7185-C and CLM-8931-PK ) as well as isotopically labelled 17 amino acids and isotopically labelled reduced glutathione (Cambridge Isotope Laboratories, MSK-A2-1.2 and CNLM-6245-10). After centrifugation for 10 min at maximum speed on a benchtop centrifuge (Eppendorf) the top, hydrophilic layer (top phase) was transferred to a new tube, dried using a nitrogen dryer and reconstituted in 20 µl water (supplemented with QReSS, Cambridge Isotope Laboratories, MSK-QRESS-KIT) by brief vortexing. Extracted metabolites were spun again and cleared supernatant was transferred to LC-MS micro vials. A small amount of each sample was pooled and serially diluted 3- and 10-fold to be used as quality controls throughout the run of each batch. I calibration curve of unlabeled T3 and T4 standards was run per experiment for concentration calculations. |