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MB Sample ID: SA288834
| Local Sample ID: | Sample_2-100_1 |
| Subject ID: | SU002847 |
| Subject Type: | Mammal |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
| Genotype Strain: | C57Bl/6 |
Select appropriate tab below to view additional metadata details:
Subject:
| Subject ID: | SU002847 |
| Subject Type: | Mammal |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
| Genotype Strain: | C57Bl/6 |
Factors:
| Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
|---|---|---|---|---|
| Sample_2-100_1 | SA288834 | FL035621 | WT C57Bl/6 | Mouse model |
| Sample_2-100_1 | SA288834 | FL035621 | CD8+ T cells | Cell Type |
| Sample_2-100_1 | SA288834 | FL035621 | 100 | [13C2-acetate] in mM |
| Sample_2-100_1 | SA288834 | FL035621 | 2 | Time (hours) |
| Sample_2-100_1 | SA288834 | FL035621 | N | ACSS2 inhibitor (Y/N) |
Collection:
| Collection ID: | CO002840 |
| Collection Summary: | CD8+ T cells and NK cells were purified by negative selection from freshly prepared mouse splenocytes (wild type vs ACSS2 knockout C57Bl/6 mice). |
| Sample Type: | Splenocytes |
Treatment:
| Treatment ID: | TR002856 |
| Treatment Summary: | Cells were incubated up to 3 hours in the in 100 or 500 µM 13C2-acetate and treated with vehicle (DMSO) or an ACSS2 inhibitor (VY-3-135). |
Sample Preparation:
| Sampleprep ID: | SP002853 |
| Sampleprep Summary: | Cells were pelleted and washed with ice-cold PBS before extraction of metabolites in 80% methanol. Samples were centrifuged at 4 °C two times (18,000 x g). Deproteinated supernatants were stored at -80 °C prior to analysis. |
Combined analysis:
| Analysis ID | AN004443 |
|---|---|
| Analysis type | MS |
| Chromatography type | HILIC |
| Chromatography system | Thermo Vanquish |
| Column | SeQuant ZIC-pHILIC (150 x 2.1mm, 5um) |
| MS Type | ESI |
| MS instrument type | Orbitrap |
| MS instrument name | Thermo Q Exactive HF-X Orbitrap |
| Ion Mode | UNSPECIFIED |
| Units | Peak Area |
Chromatography:
| Chromatography ID: | CH003338 |
| Chromatography Summary: | Hydrophilic interaction liquid chromatography (HILIC) was performed at 0.2 ml/min on a ZIC-pHILIC column (150 x 2.1 mm, 5 µM particle size, EMD Millipore) with a ZIC-pHILIC guard column (20 x 2.1 mm, EMD Millipore) at 30 °C. Solvent A was 20 mM ammonium carbonate, 0.1% ammonium hydroxide, pH 9.2, 5 µM medronic acid, and solvent B was acetonitrile. The gradient was 85% B for 2 min, 85% B to 20% B over 15 min, 20% B to 85% B over 0.1 min, and 85% B for 8.9 min. The autosampler was held at 4 °C. For each analysis, 5 µl of sample was injected. |
| Instrument Name: | Thermo Vanquish |
| Column Name: | SeQuant ZIC-pHILIC (150 x 2.1mm, 5um) |
| Column Temperature: | 30 |
| Flow Gradient: | 85% B for 2 min, 85% B to 20% B over 15 min, 20% B to 85% B over 0.1 min, and 85% B for 8.9 min |
| Flow Rate: | 0.2 ml/min |
| Solvent A: | 20 mM ammonium carbonate, 0.1% ammonium hydroxide, pH 9.2, 5 µM medronic acid |
| Solvent B: | acetonitrile |
| Chromatography Type: | HILIC |
MS:
| MS ID: | MS004190 |
| Analysis ID: | AN004443 |
| Instrument Name: | Thermo Q Exactive HF-X Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | The following parameters were used for the MS analysis: sheath gas flow rate, 30; auxiliary gas flow rate, 5; sweep gas flow rate, 0; auxiliary gas heater temperature, 200 °C; spray voltage, 3.6 kV for positive and negative modes; capillary temperature, 325 °C; and funnel RF level, 65. Samples were analyzed by full MS with polarity switching. Full MS scans were acquired at 60,000 resolution with 2 microscans and a scan range of 72-1080 m/z, automatic gain control (AGC) target of 5e6, and maximum injection time (IT) of 200 ms. |
| Ion Mode: | UNSPECIFIED |
