Return to study ST001744 main page
MB Sample ID: SA163299
Local Sample ID: | LPS13C1 |
Subject ID: | SU001821 |
Subject Type: | Cultured cells |
Subject Species: | Rattus norvegicus |
Taxonomy ID: | 10116 |
Select appropriate tab below to view additional metadata details:
Subject:
Subject ID: | SU001821 |
Subject Type: | Cultured cells |
Subject Species: | Rattus norvegicus |
Taxonomy ID: | 10116 |
Factors:
Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
---|---|---|---|---|
LPS13C1 | SA163299 | FL019445 | 1 | LPS (ug/mL) |
LPS13C1 | SA163299 | FL019445 | U-13C Glucose | Glucose |
Collection:
Collection ID: | CO001814 |
Collection Summary: | Immortalized rat astrocytes (CRL-2005) |
Sample Type: | Fibroblasts |
Treatment:
Treatment ID: | TR001834 |
Treatment Summary: | high-glucose DMEM (Gibco) containing 10% FBS and 1% penicillin/streptomycin (Gibco). Isotopic labeling was achieved through 30 min treatments with media containing 4.5 g/L U-13C-glucose (100% of the total glucose content; Cambridge Isotopes); parallel cultures were treated with 4.5 g/L natural-abundance glucose. Simultaneous to the introduction of the labeled substrate, subsets of both unlabeled and labeled cultures were treated with LPS (added to culture media to a final concentration of 1 μg/mL) for the duration of the labeling protocol. |
Sample Preparation:
Sampleprep ID: | SP001827 |
Sampleprep Summary: | Cells were then washed with phosphate buffer solution and high-performance liquid chromatography (HPLC)-grade water, quenched with 1 mL cold HPLC-grade methanol, scraped from the plate, and pelleted. Pellets were dried on a SpeedVac and subsequently lyophilized. Dried samples were weighed out and extracted with the solvent volumes adjusted to maintain a ratio of 1 mL of solvent per 1 mg of dried cellular material. The final volume of reconstitution solvent was adjusted to 100 μL per 1 mg of dried material. All cell-culture conditions (unlabeled versus labeled, control versus LPSstimulated) were sampled in triplicate. |
Combined analysis:
Analysis ID | AN002837 |
---|---|
Analysis type | MS |
Chromatography type | HILIC |
Chromatography system | Agilent 1200 |
Column | Luna Aminopropyl |
MS Type | ESI |
MS instrument type | QTOF |
MS instrument name | Agilent 6520 QTOF |
Ion Mode | NEGATIVE |
Units | peak area |
Chromatography:
Chromatography ID: | CH002100 |
Instrument Name: | Agilent 1200 |
Column Name: | Luna Aminopropyl |
Chromatography Type: | HILIC |
MS:
MS ID: | MS002630 |
Analysis ID: | AN002837 |
Instrument Name: | Agilent 6520 QTOF |
Instrument Type: | QTOF |
MS Type: | ESI |
MS Comments: | Mass spectrometry (MS) detection was carried out on an Agilent 6520 Q-TOF in negative ESI (electrospray ionization) mode with the following settings: gas temperature 325 °C, drying gas 5 L/min, nebulizer 15 psi, fragmentor 120 V, skimmer 65 V, capillary voltage −3500 V, and scan rate 1.06 spectra/s. Tandem MS (MS/MS) analyses were carried out with identical ESI parameters, and the following fragmentation and precursor ion selection settings: collision energy 10 V, precursor isolation window 1.3 amu, and scan rate 1.00 spectra/s. X13CMS R-Package was used for data processing |
Ion Mode: | NEGATIVE |