Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA042652

Local Sample ID:	18140
Subject ID:	SU000806
Subject Type:	human
Subject Species:	Homo sapiens
Taxonomy ID:	9606
Species Group:	Human

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Sample Preparation:

Sampleprep ID:	SP000813
Sampleprep Summary:	Briefly, 10 μl of undiluted plasma sample from each patient or control individual was placed in individual wells of a 96 well plate provided by the manufacturer. Samples were dried at room temperature using a nitrogen evaporator drying unit for 30 mins. Following this, 20 μl of 5% phenylisothiocyanate (PITC) reagent was added to the dried plasma spots and incubated for 20 min at RT. The plate was again dried under the nitrogen evaporator for 60 mins. Next, 300 μl of 5 mM ammonium acetate in methanol was added to each well and the mixture was incubated on a shaker (450 rpm) for 30 mins. The plate was then centrifuged at 100 g for 2 min resulting in a volume of ~ 250 μl sample in the plate (labeled as Flow injection or FIA plate). 150 μl of the sample was subsequently transferred from the FIA plate into a second plate (termed LC-MS plate). In total, all the samples were analyzed in three batches, each containing one FIA and LC-MS plate, respectively. 150 μl HPLC grade water was added to the LC-MS plate to make up the volume to 300 ul. In parallel, 500 μl of mass spectrometry injection solvent (stock solution provided by Biocrates was diluted in methanol) was added to the FIA plate. The LC-MS plate was analyzed using Multiple Reaction Monitoring post LC-MS fractionation. The FIA plate was stored at 4°C until the LC-MS plate was completed assayed.

Sampleprep ID:

SP000813

Sampleprep Summary:

Briefly, 10 μl of undiluted plasma sample from each patient or control individual was placed in individual wells of a 96 well plate provided by the manufacturer. Samples were dried at room temperature using a nitrogen evaporator drying unit for 30 mins. Following this, 20 μl of 5% phenylisothiocyanate (PITC) reagent was added to the dried plasma spots and incubated for 20 min at RT. The plate was again dried under the nitrogen evaporator for 60 mins. Next, 300 μl of 5 mM ammonium acetate in methanol was added to each well and the mixture was incubated on a shaker (450 rpm) for 30 mins. The plate was then centrifuged at 100 g for 2 min resulting in a volume of ~ 250 μl sample in the plate (labeled as Flow injection or FIA plate). 150 μl of the sample was subsequently transferred from the FIA plate into a second plate (termed LC-MS plate). In total, all the samples were analyzed in three batches, each containing one FIA and LC-MS plate, respectively. 150 μl HPLC grade water was added to the LC-MS plate to make up the volume to 300 ul. In parallel, 500 μl of mass spectrometry injection solvent (stock solution provided by Biocrates was diluted in methanol) was added to the FIA plate. The LC-MS plate was analyzed using Multiple Reaction Monitoring post LC-MS fractionation. The FIA plate was stored at 4°C until the LC-MS plate was completed assayed.