Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA052922

Local Sample ID:	CR Viv-1 24 wk -3
Subject ID:	SU000939
Subject Type:	Animal
Subject Species:	Mus musculus
Taxonomy ID:	10090
Species Group:	Mammal

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Sample Preparation:

Sampleprep ID:	SP000946
Sampleprep Summary:	Preparation of SPE on vacuum manifold: 1. Clean 60 mg Oasis HLB (Waters) spe column with 1 column volume ethyl acetate followed by 2 column volumes MeOH 2. Condition column with 2 column volume 0.1% acetic acid :5% MeOH solution 3. Close valves tightly when meniscus reaches frit over sorbent bed. Sample extraction by SPE on vacuum manifold: 4. If not in sample at collection, add 10 µL of 0.2 mg/mL of EDTA and BHT in MeOH: H2O (1:1) to sorbent bed of all columns 5. Vortex each sample a few seconds to homogenize just prior to loading. 6. Load 250µL plasma sample into SPE column. 7. Spike each column with 10µl 500nM surrogate solution 8. Load 1.5ml of 0.1% acetic acid: 5% MeOH solution to each column and let sit a few minutes. 9. Open valves to extract by gravity (use low vacuum only if necessary) 10. Wash SPE with 2 column volumes 0.1% acetic acid :5% MeOH solution 11. Pull aqueous plug from SPE with a few seconds high vacuum. 12. Further dry SPE with low vacuum. ~20min, -20psi. 13. For each sample, spike 2ml eppendorf tube with 6µl 30% glycerol in MeOH prior to eluate collection. 14. Elute spe into tube with 0.5 ml methanol followed by 1.5 mL of ethyl acetate 15. Evaporate extract until 2 µl glycerol plug is left, by Speed-vac. 16. Cap and freeze by liquid nitrogen (for shipping) or store at -80ºC until analysis.

Sampleprep ID:

SP000946

Sampleprep Summary:

Preparation of SPE on vacuum manifold: 1. Clean 60 mg Oasis HLB (Waters) spe column with 1 column volume ethyl acetate followed by 2 column volumes MeOH 2. Condition column with 2 column volume 0.1% acetic acid :5% MeOH solution 3. Close valves tightly when meniscus reaches frit over sorbent bed. Sample extraction by SPE on vacuum manifold: 4. If not in sample at collection, add 10 µL of 0.2 mg/mL of EDTA and BHT in MeOH: H2O (1:1) to sorbent bed of all columns 5. Vortex each sample a few seconds to homogenize just prior to loading. 6. Load 250µL plasma sample into SPE column. 7. Spike each column with 10µl 500nM surrogate solution 8. Load 1.5ml of 0.1% acetic acid: 5% MeOH solution to each column and let sit a few minutes. 9. Open valves to extract by gravity (use low vacuum only if necessary) 10. Wash SPE with 2 column volumes 0.1% acetic acid :5% MeOH solution 11. Pull aqueous plug from SPE with a few seconds high vacuum. 12. Further dry SPE with low vacuum. ~20min, -20psi. 13. For each sample, spike 2ml eppendorf tube with 6µl 30% glycerol in MeOH prior to eluate collection. 14. Elute spe into tube with 0.5 ml methanol followed by 1.5 mL of ethyl acetate 15. Evaporate extract until 2 µl glycerol plug is left, by Speed-vac. 16. Cap and freeze by liquid nitrogen (for shipping) or store at -80ºC until analysis.