Summary of Study ST001819

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001150. The data can be accessed directly via it's Project DOI: 10.21228/M82690 This work is supported by NIH grant, U2C- DK119886.

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Study ID	ST001819
Study Title	EC and PVC from 14-15 month-old APOE3/3, APOE3/4 and APOE4/4 mice
Study Summary	We performed a targeted lipidomic analysis on EC and PVC tissue from 14-15 month old APOE3/3, APOE3/4, and APOE4/4 mice.
Institute	Columbia University
Last Name	Nuriel
First Name	Tal
Address	630 W 168th St., P&S 12-430
Email	tn2283@cumc.columbia.edu
Phone	2123045683
Submit Date	2021-06-02
Analysis Type Detail	LC-MS
Release Date	2021-06-10
Release Version	1

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Sample Preparation:

Sampleprep ID:	SP001902
Sampleprep Summary:	Lipid and small-molecule metabolite extraction was performed using a methyl tert-butyl ether (MTBE)/methanol extraction protocol modified from previous reports (40, 41), as we have described previously (34). Briefly, individual EC or PVC tissues were homogenized in 400 ul of ice-cold methanol using a bead mill homogenizer (TissueLyser II, Qiagen) at 25 beats/sec, 2x for 45 sec each. Following homogenization, samples were incubated in 1200 ul of MTBE for 1 hr at room temperature to separate organic-soluble lipids from aqueous-soluble lipids and other small-molecules. Finally, 360 ul of ultrapure water was added (for a final ratio of 3:1:0.9 MTBE:methanol:water) to resolve the two liquid phases, and each samples were centrifuged at 10,000 x g for 10 min. For this experiment, the upper organic phase was collected from each sample and stored in a separate tube, and the remaining protein pellets were resuspended in 25 mM ammonium bicarbonate, pH 8, with 2.5% SDS. A BCA protein assay was performed on each protein fraction, and the organic phase was normalized to their protein concentration equivalent with 100% methanol. All samples were then stored at -80°C prior to analysis.

Sampleprep ID:

SP001902

Sampleprep Summary:

Lipid and small-molecule metabolite extraction was performed using a methyl tert-butyl ether (MTBE)/methanol extraction protocol modified from previous reports (40, 41), as we have described previously (34). Briefly, individual EC or PVC tissues were homogenized in 400 ul of ice-cold methanol using a bead mill homogenizer (TissueLyser II, Qiagen) at 25 beats/sec, 2x for 45 sec each. Following homogenization, samples were incubated in 1200 ul of MTBE for 1 hr at room temperature to separate organic-soluble lipids from aqueous-soluble lipids and other small-molecules. Finally, 360 ul of ultrapure water was added (for a final ratio of 3:1:0.9 MTBE:methanol:water) to resolve the two liquid phases, and each samples were centrifuged at 10,000 x g for 10 min. For this experiment, the upper organic phase was collected from each sample and stored in a separate tube, and the remaining protein pellets were resuspended in 25 mM ammonium bicarbonate, pH 8, with 2.5% SDS. A BCA protein assay was performed on each protein fraction, and the organic phase was normalized to their protein concentration equivalent with 100% methanol. All samples were then stored at -80°C prior to analysis.