Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA205976

Local Sample ID:	QC_1_J_2
Subject ID:	SU002236
Subject Type:	Cultured cells
Subject Species:	Homo sapiens
Taxonomy ID:	9606

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Treatment:

Treatment ID:	TR002248
Treatment Summary:	For lipid extraction, cell pellets were mixed with 200 µL of cold (-20°C) methanol:water (1:1, v/v) and sonicated with an ultrasonic homogenizer (UP200S, Hielscher Ultrasound Technology, HIELSCHER GmbH, Chamerau, Germany) for 16 bursts (0.5 second pulse) at 80% amplitude. Homogenates (100 µL) were mixed with 320 µL of cold (-20°C) methanol containing 1.6 ppm of sphinganine (d17:0) as the internal standard. Samples were then vortex-mixed for 2 minutes, followed by the addition of 80 µL of methyl tert-butyl ether. Subsequently, samples were vortex-mixed (1 hour, RT). After centrifugation (16,000 × g, 15°C, 10 minutes), samples were used for ultra-high performance liquid chromatography (UHPLC; Agilent 1290 Infinity II, Agilent Technologies Inc., Santa Clara, CA) coupled with (ESI) quadrupole time-of-flight (QTOF) mass spectrometry (MS) (Agilent 6546): 100 µL of each sample was divided between two UHPLC-MS vials with inserts (50 µL/each) for direct injection into the system for LC-MS analyses in positive and negative ionization modes.

Treatment ID:

TR002248

Treatment Summary:

For lipid extraction, cell pellets were mixed with 200 µL of cold (-20°C) methanol:water (1:1, v/v) and sonicated with an ultrasonic homogenizer (UP200S, Hielscher Ultrasound Technology, HIELSCHER GmbH, Chamerau, Germany) for 16 bursts (0.5 second pulse) at 80% amplitude. Homogenates (100 µL) were mixed with 320 µL of cold (-20°C) methanol containing 1.6 ppm of sphinganine (d17:0) as the internal standard. Samples were then vortex-mixed for 2 minutes, followed by the addition of 80 µL of methyl tert-butyl ether. Subsequently, samples were vortex-mixed (1 hour, RT). After centrifugation (16,000 × g, 15°C, 10 minutes), samples were used for ultra-high performance liquid chromatography (UHPLC; Agilent 1290 Infinity II, Agilent Technologies Inc., Santa Clara, CA) coupled with (ESI) quadrupole time-of-flight (QTOF) mass spectrometry (MS) (Agilent 6546): 100 µL of each sample was divided between two UHPLC-MS vials with inserts (50 µL/each) for direct injection into the system for LC-MS analyses in positive and negative ionization modes.