Metabolomics Workbench : NIH Data Repository

MB Sample ID: SA028922

Local Sample ID:	Pool_5
Subject ID:	SU000582
Subject Type:	Cells
Subject Species:	Homo sapiens
Taxonomy ID:	9606
Cell Strain Details:	Suzuki, H., Moldoveanu, Z., Hall, S., et al. IgA1-secreting cell lines from patients with IgA nephropathy produce aberrantly glycosylated IgA1. J Clin Inv. 2008, 118, 629-639
Cell Primary Immortalized:	immortalized
Cell Counts:	1x10^7 cells / pellet
Species Group:	Human

Select appropriate tab below to view additional metadata details:

Sample Preparation:

Sampleprep ID:	SP000589
Sampleprep Summary:	A total of 24 cell pellets (4 replicates from 6 cell lines) were shipped to the NIH RTI-RCMRC on dry ice and immediately stored at -80oC after being inventoried for metabolomics analysis. An addition of 500 µL of ice-cold Cell Extraction buffer (0.005 tryptophan-d5 in 50:50 Acetonitre:Water) was added to tubes containing the cell pellet samples on ice. MagNA Lyser ceramic beads (~15-20, prewashed & dried) were added to the tubes and the MagNA Lyser was used to beat samples for two 30 sec pulses at 2,000 rpm and samples were placed on a cold block for 1 min between pulses. Samples were then centrifuged at room temperature at 16,000 rcf for 4 min. A 15 mL washed conical tube was labeled Total Pool and 120 µL of each sample was added to this conical tube. There were 24 samples; thus, the resulting Total Pool was 2880 µL. The Total Pool was vortexed and 150 µL of the Total Pool was transferred to pre-labeled 1.5 mL Lo-Bind Eppendorf tubes to make two sets of 5 Total Pool samples, and 3 Equilibrium samples. The 150 uL of the remaining supernatants of each sample was transferred to a new, pre-labeled 1.5 mL Lo-Bind Eppendorf tubes and capped with disposable rubber stoppers. Samples were placed at -80 °C for 1 h and lyophilized overnight. 100 µL of 95:5 Acetonitrile:Water was added to each tube and vortexed for 2 min at 5000 rpm, followed by centrifugation at room temperature at 16,000 rcf for 4 min. The supernatants were transferred to pre-labeled autosampler vials and 3 µL was injected into SYNAPT G2-Si. All samples were prepared and analyzed in a randomized order.

Sampleprep ID:

SP000589

Sampleprep Summary:

A total of 24 cell pellets (4 replicates from 6 cell lines) were shipped to the NIH RTI-RCMRC on dry ice and immediately stored at -80oC after being inventoried for metabolomics analysis. An addition of 500 µL of ice-cold Cell Extraction buffer (0.005 tryptophan-d5 in 50:50 Acetonitre:Water) was added to tubes containing the cell pellet samples on ice. MagNA Lyser ceramic beads (~15-20, prewashed & dried) were added to the tubes and the MagNA Lyser was used to beat samples for two 30 sec pulses at 2,000 rpm and samples were placed on a cold block for 1 min between pulses. Samples were then centrifuged at room temperature at 16,000 rcf for 4 min. A 15 mL washed conical tube was labeled Total Pool and 120 µL of each sample was added to this conical tube. There were 24 samples; thus, the resulting Total Pool was 2880 µL. The Total Pool was vortexed and 150 µL of the Total Pool was transferred to pre-labeled 1.5 mL Lo-Bind Eppendorf tubes to make two sets of 5 Total Pool samples, and 3 Equilibrium samples. The 150 uL of the remaining supernatants of each sample was transferred to a new, pre-labeled 1.5 mL Lo-Bind Eppendorf tubes and capped with disposable rubber stoppers. Samples were placed at -80 °C for 1 h and lyophilized overnight. 100 µL of 95:5 Acetonitrile:Water was added to each tube and vortexed for 2 min at 5000 rpm, followed by centrifugation at room temperature at 16,000 rcf for 4 min. The supernatants were transferred to pre-labeled autosampler vials and 3 µL was injected into SYNAPT G2-Si. All samples were prepared and analyzed in a randomized order.