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MB Sample ID: SA353974
| Local Sample ID: | 210514_MDA_ZEB1_n3_2h_WT_RSL3_10_uM_21 |
| Subject ID: | SU003375 |
| Subject Type: | Cultured cells |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
Select appropriate tab below to view additional metadata details:
Subject:
| Subject ID: | SU003375 |
| Subject Type: | Cultured cells |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
Factors:
| Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
|---|---|---|---|---|
| 210514_MDA_ZEB1_n3_2h_WT_RSL3_10_uM_21 | SA353974 | FL041261 | Breast cancer cells | Sample source |
| 210514_MDA_ZEB1_n3_2h_WT_RSL3_10_uM_21 | SA353974 | FL041261 | WT | Genotype |
| 210514_MDA_ZEB1_n3_2h_WT_RSL3_10_uM_21 | SA353974 | FL041261 | RSL3 (10 µM) | Treatment |
| 210514_MDA_ZEB1_n3_2h_WT_RSL3_10_uM_21 | SA353974 | FL041261 | 2 h | Treatment Time |
Collection:
| Collection ID: | CO003368 |
| Collection Summary: | Cultured cells were washed, trypsinized, counted and flash-frozen in liquid N2 and stored at -80°C. |
| Sample Type: | Breast cancer cells |
| Storage Conditions: | -80℃ |
Treatment:
| Treatment ID: | TR003384 |
| Treatment Summary: | Brest cancer cells (MDA-MB-231 cells) for oxPE, oxPC, oxPI: Human breast cancer MDA-MB-231 wildtype (WT) cells and the stably transduced MDA-MB-231 shZeb1 (stable Zeb1 knockdown) and shCtrl cell lines (control cell line for the stable Zeb1 knockdown) (Spaderna et al. 2008, DOI: 10.1158/0008-5472.CAN-07-5682) were treated with vehicle (DMSO) or RSL3 (1 and 10 µM) for 2 h, 4 h, 6 h, 24 h or 48 h at 37°C and 5% CO2. |
Sample Preparation:
| Sampleprep ID: | SP003382 |
| Sampleprep Summary: | Phospholipids were extracted from cell pellets by successive addition of PBS pH 7.4, methanol, chloroform, and saline to a final ratio of 14:34:35:17. Evaporation of the organic layer yielded a lipid film that was dissolved in methanol and subjected to UPLC-MS/MS. |
| Extract Storage: | -80℃ |
Combined analysis:
| Analysis ID | AN005338 |
|---|---|
| Analysis type | MS |
| Chromatography type | Reversed phase |
| Chromatography system | Waters Acquity H-Class |
| Column | Waters ACQUITY UPLC BEH C8 (100 x 2.1mm,1.7um) |
| MS Type | ESI |
| MS instrument type | QTRAP |
| MS instrument name | ABI Sciex 6500+ |
| Ion Mode | NEGATIVE |
| Units | absolute intensities |
Chromatography:
| Chromatography ID: | CH004040 |
| Chromatography Summary: | Chromatographic separation of phospholipids was carried out on an Acquity BEH C8 column (1.7 μm, 2.1×100 mm, Waters, Milford, MA) using an Acquity UHPLC. |
| Instrument Name: | Waters Acquity H-Class |
| Column Name: | Waters ACQUITY UPLC BEH C8 (100 x 2.1mm,1.7um) |
| Column Temperature: | 45 |
| Flow Gradient: | The gradient was ramped from 75 to 85% B over 5 min and further increased to 100% B within 2 min, followed by isocratic elution for another 2 min. |
| Flow Rate: | 0.75 mL/min |
| Solvent A: | 90% Water, 10% Acetonitrile; 2 mM ammonium acetate |
| Solvent B: | 5% Water, 95% Acetonitrile; 2 mM ammonium acetate |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS005068 |
| Analysis ID: | AN005338 |
| Instrument Name: | ABI Sciex 6500+ |
| Instrument Type: | QTRAP |
| MS Type: | ESI |
| MS Comments: | Targeted MRM with pre-optimized settings and subsequent automated integration of selected signals using Analyst 1.6.3 or Analyst 1.7.1 (Sciex). Oxidized phospholipid species were identified by the fragmentation of [M-H]- (Ox-PE, Ox-PI) or [M+OAc]- (Ox-PC) to the saturated fatty acid anion (16:0 and 18:0) and either the PUFA anion (20:4 and 22:4) with one to three oxygen incorporated or a secondary fragment. Oxidized phospholipids were quantified based on the most intensive, specific transition to the oxidized fatty acid anions. |
| Ion Mode: | NEGATIVE |
