Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA043486

Local Sample ID:	nC18-16mar16-015
Subject ID:	SU000814
Subject Type:	Human
Subject Species:	Homo sapiens
Taxonomy ID:	9606
Species Group:	Human

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Sample Preparation:

Sampleprep ID:	SP000821
Sampleprep Summary:	The metabolite profiling as well as sample preparations will be undertaken at the Mayo Clinic Metabolomics Resource Core. The matched BM plasma and clonal PCs pellets from the stored patient samples will undergo metabolite extraction and analysis by LC-MS. They will be deproteinized with cold methanol (1:4 ratio) and centrifuged at 15,000 g for 20 minutes at 4οC. The supernatants will be divided into 2 aliquots and dried down under a stream of nitrogen for 90 minutes and stored at -20C until ready for analysis. These samples will undergo preparation for metabolite separation and analysis on a Time-of-Flight (TOF) Mass Spectrometer (Agilent Technologies 6220 TOF) coupled with an Ultra High Pressure Liquid Chromatograph (Waters Acquity). The dried metabolites will be re‐suspended in H2O/acetonitrile solution containing injection standards. Profiling data will be acquired under both positive and negative electrospray ionization modes separately over a scan range of 50 - 1200 m/z at a resolution of 10,000. Metabolite separation will be achieved using two columns of differing polarity, a hydrophilic interaction column (HILIC, ethylene-bridged hybrid 2.1 x 150 mm, 1.7 mm; Waters) and a reversed-phase C18 column (high-strength silica 2.1 x 150 mm, 1.8 mm; Waters). For positive ion detection, mobile phase A will contain 100% water with 0.1% formic acid and mobile phase B will contain 100% methanol with 0.1% formic acid. For negative ion detection, formic acid will be replaced with 0.1% ammonium bicarbonate. For each column, the run time will be 20 min using a flow rate of 400 microL/min. A total of four runs per sample will be performed to give maximum coverage of metabolites. Samples will be injected in triplicate with blank injections between samples. Quality control samples, made up of a subset of samples from the study will be injected several times during a run.

Sampleprep ID:

SP000821

Sampleprep Summary:

The metabolite profiling as well as sample preparations will be undertaken at the Mayo Clinic Metabolomics Resource Core. The matched BM plasma and clonal PCs pellets from the stored patient samples will undergo metabolite extraction and analysis by LC-MS. They will be deproteinized with cold methanol (1:4 ratio) and centrifuged at 15,000 g for 20 minutes at 4οC. The supernatants will be divided into 2 aliquots and dried down under a stream of nitrogen for 90 minutes and stored at -20C until ready for analysis. These samples will undergo preparation for metabolite separation and analysis on a Time-of-Flight (TOF) Mass Spectrometer (Agilent Technologies 6220 TOF) coupled with an Ultra High Pressure Liquid Chromatograph (Waters Acquity). The dried metabolites will be re‐suspended in H2O/acetonitrile solution containing injection standards. Profiling data will be acquired under both positive and negative electrospray ionization modes separately over a scan range of 50 - 1200 m/z at a resolution of 10,000. Metabolite separation will be achieved using two columns of differing polarity, a hydrophilic interaction column (HILIC, ethylene-bridged hybrid 2.1 x 150 mm, 1.7 mm; Waters) and a reversed-phase C18 column (high-strength silica 2.1 x 150 mm, 1.8 mm; Waters). For positive ion detection, mobile phase A will contain 100% water with 0.1% formic acid and mobile phase B will contain 100% methanol with 0.1% formic acid. For negative ion detection, formic acid will be replaced with 0.1% ammonium bicarbonate. For each column, the run time will be 20 min using a flow rate of 400 microL/min. A total of four runs per sample will be performed to give maximum coverage of metabolites. Samples will be injected in triplicate with blank injections between samples. Quality control samples, made up of a subset of samples from the study will be injected several times during a run.