Return to study ST002713 main page
MB Sample ID: SA273540
| Local Sample ID: | A375 VR-Rano 1 |
| Subject ID: | SU002818 |
| Subject Type: | Cultured cells |
| Subject Species: | Homo sapiens |
| Gender: | Female |
| Cell Biosource Or Supplier: | ATCC |
| Cell Strain Details: | A375 |
Select appropriate tab below to view additional metadata details:
Subject:
| Subject ID: | SU002818 |
| Subject Type: | Cultured cells |
| Subject Species: | Homo sapiens |
| Gender: | Female |
| Cell Biosource Or Supplier: | ATCC |
| Cell Strain Details: | A375 |
Factors:
| Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
|---|---|---|---|---|
| A375 VR-Rano 1 | SA273540 | FL035353 | VEMURA+RANO | Treatment |
Collection:
| Collection ID: | CO002811 |
| Collection Summary: | All cell lines had been authenticated in 2021. Cells were expanded to generate enough vials from a single batch before the start of the study. Cell lines were cultured in Dulbecco’s Eagle’s Medium (DMEM) (cat#61965026, Gibco) supplemented with 10% fetal bovine serum (cat#10500064, Gibco) plus 1% penicillin/streptomycin (cat#15140122, Gibco). Cells were grown at 37°C in a 5% CO2 environment. |
| Sample Type: | Melanoma cells |
Treatment:
| Treatment ID: | TR002827 |
| Treatment Summary: | Melanoma cells seeded in 6-well plates were treated for 7 days with a high dose (5 or 10μM as indicated) of vemurafenib before switching to a lower concentration of the drug (0,5 or 1μM). Fresh medium and vemurafenib was added once weekly for further 3-4 weeks until arising colonies grew to confluence. Fatty Acid oxidation inhibitors ranolazine, etomoxir or thiroridazine were added once a week. |
Sample Preparation:
| Sampleprep ID: | SP002824 |
| Sampleprep Summary: | Lipids from frozen cell pellets were extracted with cold methanol at a concentration of 2 million cells/mL. Samples were vortexed 30 min at 4 degrees C then supernatants clarified by centrifugation (10 min, 10,000 g, 4 degrees C) and transferred to autosampler vials. |
| Processing Storage Conditions: | 4℃ |
| Extract Storage: | -80℃ |
Combined analysis:
| Analysis ID | AN004397 | AN004398 |
|---|---|---|
| Analysis type | MS | MS |
| Chromatography type | Reversed phase | Reversed phase |
| Chromatography system | Thermo Vanquish | Thermo Vanquish |
| Column | Phenomenex Kinetex C18 (30 x 2.1mm,1.7um) | Phenomenex Kinetex C18 (30 x 2.1mm,1.7um) |
| MS Type | ESI | ESI |
| MS instrument type | Orbitrap | Orbitrap |
| MS instrument name | Thermo Q Exactive Orbitrap | Thermo Q Exactive Orbitrap |
| Ion Mode | NEGATIVE | POSITIVE |
| Units | peak area | peak area |
Chromatography:
| Chromatography ID: | CH003299 |
| Chromatography Summary: | Negative C18 |
| Instrument Name: | Thermo Vanquish |
| Column Name: | Phenomenex Kinetex C18 (30 x 2.1mm,1.7um) |
| Column Temperature: | 45 |
| Flow Gradient: | 0-3 min 10-95% B at 0.3 mL/min, 3-4.2 min hold at 95% B at 0.3 mL/min, 4.2-4.3 min drop to 10% B at 0.45 mL/min, 4.3-4.9 min hold at 10%B while lowering flow rate to 0.4 mL/min, 4.9-5 min hold at 10%B while lowering flow rate to 0.3 mL/min. |
| Flow Rate: | see gradient |
| Sample Injection: | 10 uL |
| Solvent A: | 75% water 25% acetonitrile 5 mM ammonium acetate |
| Solvent B: | 90% isopropanol 10% acetonitrile 5 mM ammonium acetate |
| Chromatography Type: | Reversed phase |
| Chromatography ID: | CH003300 |
| Chromatography Summary: | Positive C18 |
| Instrument Name: | Thermo Vanquish |
| Column Name: | Phenomenex Kinetex C18 (30 x 2.1mm,1.7um) |
| Column Temperature: | 45 |
| Flow Gradient: | 0-3 min 30-100% B at 0.3 mL/min, 3-4.2 min hold at 100% B at 0.3 mL/min, 4.2-4.3 min 100-30% B at 0.4 mL/min, 4.3-4.9 min hold at 30%B and 0.4 mL/min, 4.9-5 min hold at 30%B while lowering flow rate to 0.3 mL/min |
| Flow Rate: | see gradient |
| Sample Injection: | 10 uL |
| Solvent A: | 75% water 25% acetonitrile 5 mM ammonium acetate |
| Solvent B: | 90% isopropanol 10% acetonitrile 5 mM ammonium acetate |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS004146 |
| Analysis ID: | AN004397 |
| Instrument Name: | Thermo Q Exactive Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | The Q Exactive was run independently in positive and negative ion mode, scanning using full MS from 125-1500 m/z at 70,000 resolution and top 10 data-dependent MS2 at 17,500 resolution. Electrospray ionization was achieved with 45 Arb sheath gas, 25 Arb auxiliary gas, and 4 kV spray voltage. Calibration was performed prior to the run using the PierceTM Positive and Negative Ion Calibration Solutions (Thermo Fisher Scientific). Run order of samples was randomized and technical replicates were injected after every 4 samples to assess quality control. |
| Ion Mode: | NEGATIVE |
| MS ID: | MS004147 |
| Analysis ID: | AN004398 |
| Instrument Name: | Thermo Q Exactive Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | The Q Exactive was run independently in positive and negative ion mode, scanning using full MS from 125-1500 m/z at 70,000 resolution and top 10 data-dependent MS2 at 17,500 resolution. Electrospray ionization was achieved with 45 Arb sheath gas, 25 Arb auxiliary gas, and 4 kV spray voltage. Calibration was performed prior to the run using the PierceTM Positive and Negative Ion Calibration Solutions (Thermo Fisher Scientific). Run order of samples was randomized and technical replicates were injected after every 4 samples to assess quality control. |
| Ion Mode: | POSITIVE |
