Summary of Study ST000246
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 PR000125. The data can be accessed directly via it's Project DOI: 10.21228/M8DW2B This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
| Study ID | ST000246 |
| Study Title | Lipid Extraction Efficiency Comparison |
| Study Type | Lipid Extraction Efficiency Comparison |
| Study Summary | Aliquots of Jurkat T-lymphocyte cells were extracted using the Folch or Matyash method at different total volumes and run in triplicate to calculate the extraction efficiency for each method. |
| Institute | University of Florida |
| Department | Dept. of Chemistry/SECIM |
| Laboratory | Biomedical Mass Spectrometry Lab |
| Last Name | Ulmer |
| First Name | Candice |
| Address | R3-226 Academic Research Building, Department of Biochemistry and Molecular Biology, PO Box 100245, Gainesville, FL 32610-0245 |
| czulmer@chem.ufl.edu | |
| Phone | (352) 392-0515 |
| Submit Date | 2015-09-15 |
| Num Groups | 4 |
| Total Subjects | 12 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | raw(Thermo) |
| Uploaded File Size | 2.2 G |
| Analysis Type Detail | LC-MS |
| Release Date | 2016-12-22 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR000125 |
| Project DOI: | doi: 10.21228/M8DW2B |
| Project Title: | Mammalian Suspension-Cultured Cellular Metabolomics Workflow |
| Project Type: | Metabolomics, Lipidomics, Untargeted |
| Project Summary: | A workflow was optimized for the sample preparation of a single suspension-cultured cell pellet for both metabolomics and lipidomics analysis. Jurkat T-lymphocyte cells were washed with various rinsing solutions and the lipids extracted using different lipid extraction protocols to allow for the most reproducible and quantitative method. |
| Institute: | University of Florida |
| Department: | Dept. of Chemistry/SECIM |
| Laboratory: | Biomedical Mass Spectrometry Lab |
| Last Name: | Yost |
| First Name: | Richard |
| Address: | P.O. Box 117200, Gainesville, FL 32611 |
| Email: | ryost@aa.ufl.edu |
| Phone: | (352) 392-0515 |
| Funding Source: | JDRF Research Grant |
Subject:
| Subject ID: | SU000266 |
| Subject Type: | Human cells |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
| Cell Biosource Or Supplier: | ATCC - Supplier |
| Subject Comments: | P?+10 |
| Cell Primary Immortalized: | immortalized |
| Cell Passage Number: | P?+10 |
| Cell Counts: | P?+10 |
| Species Group: | Mammals |
Factors:
Subject type: Human cells; Subject species: Homo sapiens (Factor headings shown in green)
| mb_sample_id | local_sample_id | Extraction Method |
|---|---|---|
| SA011335 | folch_low | Folch |
| SA011336 | folch_high | Folch |
| SA011337 | mtbe_high | Matyash |
| SA011338 | mtbe_low | Matyash |
| Showing results 1 to 4 of 4 |
Collection:
| Collection ID: | CO000257 |
| Collection Summary: | - |
| Sample Type: | Cells - Jurkat, Clone E6-1 |
| Collection Method: | Centrifugation |
| Collection Frequency: | 1200 rpm |
| Collection Duration: | 5 min |
| Volumeoramount Collected: | 10 mL of cell suspension |
| Storage Conditions: | -80°C |
| Tissue Cell Quantity Taken: | 1e6 cells |
Treatment:
| Treatment ID: | TR000277 |
| Cell Storage: | Liquid Nitrogen, 5% DMSO |
| Cell Growth Container: | 10 cm2 culture dish |
| Cell Media: | RPMI 1640 |
| Cell Envir Cond: | 95% humidity, 5% CO2 |
| Cell Harvesting: | centrifugation |
Sample Preparation:
| Sampleprep ID: | SP000271 |
| Sampleprep Summary: | Cells were extracted using either the Folch or Matyash method. The total volume of MeOH in each extraction method was varied (0.5 mL or 2.0 mL) while the ratio of MeOH to the other solvents was maintained. |
| Sampleprep Protocol Filename: | SOP_LipidExtractionProtocol.pdf |
| Processing Method: | cell lysis |
| Processing Storage Conditions: | on ice |
| Extraction Method: | Folch or Matyash lipid extraction method |
| Extract Concentration Dilution: | 1e6 cells were extracted with the solvent systems explained in the Sample Prep Summary |
| Sample Resuspension: | 100 μL of IPA |
| Sample Spiking: | internal standards: lysophosphatidylcholine (LPC 17:0 and LPC 19:0), phosphatidylcholine (PC 17:0/17:0 and PC 19:0/19:0), phosphatidylethanolamine (PE 15:0/15:0 and PE 17:0/17:0), phosphatidylserine (PS 14:0/14:0 and PS 17:0/17:0), and phosphatidylglycerol (PG 14:0/14:0 and PG 17:0/17:0), triacylglyceride (TG 15:0/15:0/15:0 and TG 17:0/17:0/17:0) |
| Cell Type: | Jurkat, Clone E6-1 |
Combined analysis:
| Analysis ID | AN000384 |
|---|---|
| Analysis type | MS |
| Chromatography type | Reversed phase |
| Chromatography system | Thermo Dionex Ultimate 3000 RS |
| Column | Supelco Analytical Titan C18 |
| MS Type | ESI |
| MS instrument type | Orbitrap |
| MS instrument name | Thermo Q Exactive Orbitrap |
| Ion Mode | POSITIVE |
| Units | Peak Area |
Chromatography:
| Chromatography ID: | CH000276 |
| Chromatography Summary: | Reversed Phase |
| Methods Filename: | m_lipid_pos_switch_100 |
| Instrument Name: | Thermo Dionex Ultimate 3000 RS |
| Column Name: | Supelco Analytical Titan C18 |
| Column Pressure: | 1241 bar (max) |
| Column Temperature: | 30 C |
| Flow Gradient: | 32% B at 0 min, 40% B at 1 min, a hold at 40% B until 1.5 min, 45% B at 4 min, 50% B at 5 min, 60% B at 8 min, 70% B at 11 min, and 80% B at 14 min |
| Flow Rate: | 0.5 mL/min |
| Injection Temperature: | 4 C |
| Sample Injection: | 2 μL |
| Solvent A: | 40% water/60% acetonitrile; 0.1% formic acid; 10 mM ammonium formate |
| Solvent B: | 90% isopropanol/8% acetonitrile/2% water; 0.1% formic acid; 10 mM ammonium formate |
| Analytical Time: | 21 min |
| Randomization Order: | 3,10,12,11,8,6,5,4,7,9,2,1 |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS000330 |
| Analysis ID: | AN000384 |
| Instrument Name: | Thermo Q Exactive Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| Ion Mode: | POSITIVE |
| Capillary Temperature: | 320 |
| Dry Gas Flow: | 30 |
| Mass Accuracy: | <2 ppm |
| Spray Voltage: | 3.5 kV |
| Desolvation Gas Flow: | .raw |
| Scan Range Moverz: | 100-1500 |
| Skimmer Voltage: | 15 V |
| Acquisition Parameters File: | QE1_CZU_08_sequence |
