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MB Sample ID: SA200933
| Local Sample ID: | CON-R-8RL |
| Subject ID: | SU002177 |
| Subject Type: | Mammal |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
| Age Or Age Range: | 21 weeks |
| Gender: | Male |
Select appropriate tab below to view additional metadata details:
Subject:
| Subject ID: | SU002177 |
| Subject Type: | Mammal |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
| Age Or Age Range: | 21 weeks |
| Gender: | Male |
Factors:
| Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
|---|---|---|---|---|
| CON-R-8RL | SA200933 | FL024375 | Control | Treatment |
Collection:
| Collection ID: | CO002170 |
| Collection Summary: | C57BL/6J male mice were purchased from Jackson Laboratories. At 6 weeks of age, mice were fed lean control (10% kcal from fat) or high-fat diet (60% kcal from fat) for 15 weeks (Research Diets, New Brunswick, NJ). Ob/ob mice, a genetic model of obesity driven by leptin deficiency, were purchased from Jackson laboratories at age 10 weeks and were fed a normal chow diet for an additional 5 weeks. |
| Sample Type: | Lung |
Treatment:
| Treatment ID: | TR002189 |
| Treatment Summary: | C57BL/6J male mice were purchased from Jackson Laboratories. At 6 weeks of age, mice were fed lean control (10% kcal from fat) or high-fat diet (60% kcal from fat) for 15 weeks (Research Diets, New Brunswick, NJ). Ob/ob mice, a genetic model of obesity driven by leptin deficiency, were purchased from Jackson laboratories at age 10 weeks and were fed a normal chow diet for an additional 5 weeks. |
Sample Preparation:
| Sampleprep ID: | SP002183 |
| Sampleprep Summary: | Mouse lungs were weighed into Eppendorf tubes. The lung tissues were mashed using a clean metal spatula prior to extraction. The samples were extracted using a liquid-liquid partition with water (250 µL), methanol (300 µL), and methyl tert-butyl ether (MTBE). Avanti’s deuterated lipid mix, Equisplash, was used as an internal standard. The deuterated mix was spiked into the methanol at 1.5 µg/mL and used for extraction. The extracts were centrifuged at 20,000 rcf for 10 minutes to facilitate phase separation. The top layer was removed, dried, and reconstituted in 150 µL of isopropanol for analysis. |
| Processing Storage Conditions: | -80℃ |
| Sample Resuspension: | 150 µL of IPA |
Combined analysis:
| Analysis ID | AN003419 |
|---|---|
| Analysis type | MS |
| Chromatography type | Reversed phase |
| Chromatography system | Waters Acquity I-Class |
| Column | Waters Acquity BEH C18 (100 x 2mm,1.7um) |
| MS Type | ESI |
| MS instrument type | Orbitrap |
| MS instrument name | Thermo Q Exactive HF-X Orbitrap |
| Ion Mode | POSITIVE |
| Units | Peak Area |
Chromatography:
| Chromatography ID: | CH002529 |
| Instrument Name: | Waters Acquity I-Class |
| Column Name: | Waters Acquity BEH C18 (100 x 2mm,1.7um) |
| Column Temperature: | 45 |
| Flow Gradient: | Starting composition was 32% B, which increased to 40% B at 1 min (held until 1.5 min) then 45% B at 4 minutes. This was increased to 50% B at 5 min, 60% B at 8 min, 70% B at 11 min, and 80% B at 14 min (held until 16 min). At 16 min the composition switched back to starting conditions (32% B) and was held for 4 min to re-equilibrate the column. |
| Flow Rate: | 0.2 mL/min |
| Injection Temperature: | 10 |
| Solvent A: | 40% water/60% acetonitrile; 10 mM ammonium formate |
| Solvent B: | 90% isopropanol/10% acetonitrile; 10 mM ammonium formate |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS003184 |
| Analysis ID: | AN003419 |
| Instrument Name: | Thermo Q Exactive HF-X Orbitrap |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | Samples were analyzed with positive/negative ionization switching. 60,000 resolution. Top 5 DDA. |
| Ion Mode: | POSITIVE |
| Collision Energy: | 25, 35, 45 V stepped collision energy |
| Fragmentation Method: | HCD |
| Ionization: | HESI |
| Mass Accuracy: | 5 ppm |
