Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA235508

Local Sample ID:	FP11b-3-PE
Subject ID:	SU002435
Subject Type:	Cultured cells
Subject Species:	Homo sapiens
Taxonomy ID:	9606/10090

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Sample Preparation:

Sampleprep ID:	SP002441
Sampleprep Summary:	Cells were detached by incubation with trypsin and suspended in PBS. Protein concentration was determined by the bicinchonic acid method (Thermo Fisher Scientific, Rockford, IL). Total lipids were extracted from 50 μg of cellular protein by the Bligh and Dyer method. Cells were dissolved in methanol/chloroform/water at 2:1:0.8 v/v/v and then 50 pmol of 1,2-adidodecanoyl-sn-glycero-3-phosphocholine (Avanti Polar Lipids, Alabaster, AL) and 1,2-didodecanoyl-sn-glycero-3-phosphoethanolamine (Avanti Polar Lipids) were added as internal standards. After incubation for 5 min at room temperature, 1 ml each of water and chloroform were added and the samples were then centrifuged at 2,000 rpm for 5 min in Himac CF-16RX (Hitachi Koki, Tokyo, Japan) to collect the lower organic phase. To re-extract the lipids from the water phase, 1 ml chloroform was added. The combined organic phase was evaporated under a nitrogen stream and the extracted lipids were dissolved in methanol.

Sampleprep ID:

SP002441

Sampleprep Summary:

Cells were detached by incubation with trypsin and suspended in PBS. Protein concentration was determined by the bicinchonic acid method (Thermo Fisher Scientific, Rockford, IL). Total lipids were extracted from 50 μg of cellular protein by the Bligh and Dyer method. Cells were dissolved in methanol/chloroform/water at 2:1:0.8 v/v/v and then 50 pmol of 1,2-adidodecanoyl-sn-glycero-3-phosphocholine (Avanti Polar Lipids, Alabaster, AL) and 1,2-didodecanoyl-sn-glycero-3-phosphoethanolamine (Avanti Polar Lipids) were added as internal standards. After incubation for 5 min at room temperature, 1 ml each of water and chloroform were added and the samples were then centrifuged at 2,000 rpm for 5 min in Himac CF-16RX (Hitachi Koki, Tokyo, Japan) to collect the lower organic phase. To re-extract the lipids from the water phase, 1 ml chloroform was added. The combined organic phase was evaporated under a nitrogen stream and the extracted lipids were dissolved in methanol.