Please read Metabolite Processing Precautions before proceeding.
⦁ Prepare and separate all material and solutions before you start. Check the Recommended Resources List.
⦁ Allow serum/plasma samples to thaw on ice at 4°C for 30–60 min.
⦁ While samples are thawing, prepare one tube labeled as QC (pooled quality control). NOTE: In this tube you will prepare one aliquot comprised of all your samples (we recommend taking from each sample 10% of the volume used in the extraction). From that, make sure to pick up a tube to contain the final volume. Ex: For extraction of 24 plasma samples you need a tube able to fit 12 ml, 9.6 ml of extraction solution and 2.4 mL of combined samples (pool of 10 µL from each sample).
⦁ Vortex and transfer 100 µL of each sample to a new 2 mL tube containing 400 µL of ice cold extraction solution A.
⦁ Transfer 10 µL of each sample to the QC tube containing ice cold extraction solution A (Keep a 1:4 vol/vol ratio).
⦁ Vortex for 15 s and pelletize the protein precipitate in a centrifuge operating at 4°C and at 15,800 g for 15 min.
⦁ Transfer supernatant to a new labeled 2.0-ml tube and freeze-dry the samples in a lypholyzer. PAUSE POINT: Store the samples at -20°C for up to 12 weeks.
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