Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA064556

Local Sample ID:	sgc_050516r_HL60_Zn1h_T_01
Subject ID:	SU001068
Subject Type:	Cultured cells
Subject Species:	Homo sapiens
Taxonomy ID:	9606
Gender:	Female
Cell Biosource Or Supplier:	ATCC
Cell Primary Immortalized:	Yes
Cell Passage Number:	12
Cell Counts:	1 million/mL
Species Group:	human

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Sample Preparation:

Sampleprep ID:	SP001075
Sampleprep Summary:	The remaining HL60 cells (approximately one million) were lysed in 100 µL of 1:1:2 MeCNCH3CN: CH3MeOH:NH4HCO3 (50 mM, pH 8.0) followed by one freeze-thaw cycle (three minutes each) and a ten-minute sonication in an ice bath. Protein concentration was obtained using the Pierce BCA Protein Assay Kit (Thermo Fisher Scientific). An aliquot of 100 or 150 µg of each sample in a total of 100 µL of lysis buffer was precipitated with 300 µL of ice cold 75:25 acetone:methanol for 2 h at -80°C. Samples were spun for 15 minutes at 6,800 x g and the supernatant was removed and utilized for metabolomics analysis. The pellets were rinsed with 300 µL of ice cold acetone and spun as above. Acetone was removed and the pellet was allowed to dry briefly. Metabolite supernatants were dried and reconstituted in 50 uL of appropriate reverse phase liquid chromatography (0.1% formic acid in 98:2 H2O: CH3CNMeCN) or hydrophilic interaction chromatography (80:20 CH3CNMeCN: H2O) compatible buffers prior to analyses.

Sampleprep ID:

SP001075

Sampleprep Summary:

The remaining HL60 cells (approximately one million) were lysed in 100 µL of 1:1:2 MeCNCH3CN: CH3MeOH:NH4HCO3 (50 mM, pH 8.0) followed by one freeze-thaw cycle (three minutes each) and a ten-minute sonication in an ice bath. Protein concentration was obtained using the Pierce BCA Protein Assay Kit (Thermo Fisher Scientific). An aliquot of 100 or 150 µg of each sample in a total of 100 µL of lysis buffer was precipitated with 300 µL of ice cold 75:25 acetone:methanol for 2 h at -80°C. Samples were spun for 15 minutes at 6,800 x g and the supernatant was removed and utilized for metabolomics analysis. The pellets were rinsed with 300 µL of ice cold acetone and spun as above. Acetone was removed and the pellet was allowed to dry briefly. Metabolite supernatants were dried and reconstituted in 50 uL of appropriate reverse phase liquid chromatography (0.1% formic acid in 98:2 H2O: CH3CNMeCN) or hydrophilic interaction chromatography (80:20 CH3CNMeCN: H2O) compatible buffers prior to analyses.