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MB Sample ID: SA177874
| Local Sample ID: | MDAMB231_5_3 |
| Subject ID: | SU001998 |
| Subject Type: | Cultured cells |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
Select appropriate tab below to view additional metadata details:
Subject:
| Subject ID: | SU001998 |
| 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 |
|---|---|---|---|---|
| MDAMB231_5_3 | SA177874 | FL022024 | Factors;Control | Factors |
Collection:
| Collection ID: | CO001991 |
| Collection Summary: | The cells were seeded at a density of 1 × 10^6 cells/mL in a culture flask. The cells which had grown to 80-90% confluence, were harvested by treatment with trypsin- ethylenediaminetetraacetic acid and centrifuged. To remove the residual medium, the cell pellet was washed with 1× phosphate buffered saline twice. The cells were freeze-dried and stored at −70 °C until further analysis. |
| Sample Type: | Breast cancer cells |
| Storage Conditions: | Described in summary |
Treatment:
| Treatment ID: | TR002010 |
| Treatment Summary: | Cells at 70–80% confluency were subjected to irradiation using a ^60CO Theratron-780 teletherapy unit at a dose rate of 1.52 Gy per minute. The cells were subjected to 25 cycles of 2 Gy irradiation over 5 weeks, and the surviving cells were designated as MDA-MB-231/RR cells. |
| Cell Media: | DMEM supplemented with 10% FBS and 1% penicillin-streptomycin |
| Cell Envir Cond: | A humidified incubator at 37 °C (95% air and 5% CO2) |
Sample Preparation:
| Sampleprep ID: | SP002004 |
| Sampleprep Summary: | Extraction of metabolites and intact lipid species was performed using a modified Folch procedur. Briefly, 1 mL of ice-cold chloroform containing 0.1% BHT and 0.5 mL of ice-cold methanol containing 0.1% BHT were added to the freeze-dried cells and vortexed. The mixture were sonicated for 30 min at 4 °C and incubated for 40 min on ice with shaking. Ice-cold water containing 0.1% BHT (0.38 mL) was added to the mixture for phase separation, and then centrifuged. The upper (methanol) and lower (chloroform) phases were dried under nitrogen gas and used GC-MS and nanoESI-MS analyses, respectively. To conduct metabolic profiling using GC-MS, derivatization reaction was conducted by adding 30 μL of 20,000 μg/mL methoxylamine hydrochloride in pyridine, 10 μL of myristic-d27 acid (300 μg/mL as an internal standard), and 50 μL of BSTFA containing 1% TMCS into dried (methanol phase) sample and incubated for 60 min at 65 °C. After derivatization, 15 μL of each sample was pooled for quality control (QC) and analyzed in sextuplicate. To conduct metabolic profiling using nanoESI-MS, dried (chloroform phase) sample was resuspended with 130 μL of buffer solution (7.5 mM ammonium acetate in methanol-chloroform (9:1, v/v)) and 10 μL of each resuspended sample was pooled for QC and analyzed in sextuplicate. |
| Processing Storage Conditions: | Described in summary |
| Extraction Method: | A modified Folch method |
| Extract Storage: | -80℃ |
| Sample Resuspension: | Described in summary |
| Sample Derivatization: | Described in summary |
Combined analysis:
| Analysis ID | AN003119 | AN003120 | AN003121 |
|---|---|---|---|
| Analysis type | MS | MS | MS |
| Chromatography type | GC | None (Direct infusion) | None (Direct infusion) |
| Chromatography system | Agilent 7890A | automated nanoinfusion/nanospray source | automated nanoinfusion/nanospray source |
| Column | Agilent DB5-MS (30m x 0.25mm, 0.25um) | None | None |
| MS Type | EI | ESI | ESI |
| MS instrument type | Triple axis detector | Ion trap | Ion trap |
| MS instrument name | Agilent 5975C | Thermo LTQ XL | Thermo LTQ XL |
| Ion Mode | POSITIVE | POSITIVE | NEGATIVE |
| Units | relative intensity/µg proteins | relative intensity/µg proteins | relative intensity/µg proteins |
Chromatography:
| Chromatography ID: | CH002304 |
| Instrument Name: | Agilent 7890A |
| Column Name: | Agilent DB5-MS (30m x 0.25mm, 0.25um) |
| Internal Standard: | myristic-d27 acid (300 μg/mL as an internal standard |
| Chromatography Type: | GC |
| Chromatography ID: | CH002305 |
| Instrument Name: | automated nanoinfusion/nanospray source |
| Column Name: | None |
| Chromatography Type: | None (Direct infusion) |
| Chromatography ID: | CH002306 |
| Instrument Name: | automated nanoinfusion/nanospray source |
| Column Name: | None |
| Chromatography Type: | None (Direct infusion) |
MS:
| MS ID: | MS002900 |
| Analysis ID: | AN003119 |
| Instrument Name: | Agilent 5975C |
| Instrument Type: | Triple axis detector |
| MS Type: | EI |
| MS Comments: | The initial oven temperature was set at 70 °C and then increased to 190°C (5 °C/min), 240 °C (6 °C/min), and 280 °C (5 °C/min). Data was processed using Expressionist® MSX software (version 2013.0.39, Genedata, Basel, Switzerland). |
| Ion Mode: | POSITIVE |
| MS ID: | MS002901 |
| Analysis ID: | AN003120 |
| Instrument Name: | Thermo LTQ XL |
| Instrument Type: | Ion trap |
| MS Type: | ESI |
| MS Comments: | The raw data files (*.raw) were transformed to *.mzXML using the ProteoWizard MSConvert. Data was processed using Expressionist® MSX software (version 2013.0.39, Genedata, Basel, Switzerland). |
| Ion Mode: | POSITIVE |
| Capillary Temperature: | 200 °C |
| Capillary Voltage: | 20 V |
| Gas Pressure: | 0.4 psi |
| Ion Spray Voltage: | 1.4 kV |
| MS ID: | MS002902 |
| Analysis ID: | AN003121 |
| Instrument Name: | Thermo LTQ XL |
| Instrument Type: | Ion trap |
| MS Type: | ESI |
| MS Comments: | The raw data files (*.raw) were transformed to *.mzXML using the ProteoWizard MSConvert. Data was processed using Expressionist® MSX software (version 2013.0.39, Genedata, Basel, Switzerland). |
| Ion Mode: | NEGATIVE |
| Capillary Temperature: | 200 °C |
| Capillary Voltage: | −2 V |
| Gas Pressure: | 0.6 psi |
| Ion Spray Voltage: | 1.7 kV |
