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MB Sample ID: SA235375
| Local Sample ID: | Naive_248 |
| Subject ID: | SU002434 |
| Subject Type: | Mammal |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
| Age Or Age Range: | 12-24 weeks |
| Gender: | Male |
| Animal Animal Supplier: | Jackson |
| Animal Light Cycle: | 12L/12D |
Select appropriate tab below to view additional metadata details:
Subject:
| Subject ID: | SU002434 |
| Subject Type: | Mammal |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
| Age Or Age Range: | 12-24 weeks |
| Gender: | Male |
| Animal Animal Supplier: | Jackson |
| Animal Light Cycle: | 12L/12D |
Factors:
| Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
|---|---|---|---|---|
| Naive_248 | SA235375 | FL029196 | Naïve | Group |
Collection:
| Collection ID: | CO002427 |
| Collection Summary: | Whole blood was extracted from animals at the time of euthanization from the heart chamber. Blood was collected into blood collection tubes (Fisher Scientific; #02-675-185) and spun for 10 min at 11,000g. Serum was collected and frozen in liquid nitrogen. 25uL of plasma was extracted with 500uL of acetonitrile by vortexing and centrifugation at 10min at 14,000rpm. 450uL of supernatant was transferred to new tube and dried via SpeedVac. Dried samples were reconstituted with 25uL of 50% methanol and transferred to autosampler vials. Injection volume =10uL in PRM mode for detection and quantification of 10 different analytes. Metabolite mixture was analyzed on Thermo Orbitrap IDX MS system coupled to a Vanquish UPLC system. Samples were transported via the autosampler (10uL injection volume) onto a Waters BEH C18 column. Runtime was 15min in PRM mode. Buffer A: 0.1% formic acid in water. Buffer B: 0.1% formic acid in methanol. LC Gradient: 0min: 0% B, 8min: 50% B, 9 min: 98% B, 13min: 98% B. Recalibration of system up to 15 min at 0% B for next injection. |
| Sample Type: | Blood (serum) |
| Collection Method: | Cardiac Puncture |
| Collection Location: | Heart |
| Storage Conditions: | Described in summary |
Treatment:
| Treatment ID: | TR002446 |
| Treatment Summary: | Mice were subjected to 3weeks of unpredictable chronic mild restraint stress or kept in a naive setting |
Sample Preparation:
| Sampleprep ID: | SP002440 |
| Sampleprep Summary: | Whole blood was extracted from animals at the time of euthanization from the heart chamber. Blood was collected into blood collection tubes (Fisher Scientific; #02-675-185) and spun for 10 min at 11,000g. Serum was collected and frozen in liquid nitrogen. 25uL of plasma was extracted with 500uL of acetonitrile by vortexing and centrifugation at 10min at 14,000rpm. 450uL of supernatant was transferred to new tube and dried via SpeedVac. Dried samples were reconstituted with 25uL of 50% methanol and transferred to autosampler vials. Injection volume =10uL in PRM mode for detection and quantification of 10 different analytes. Metabolite mixture was analyzed on Thermo Orbitrap IDX MS system coupled to a Vanquish UPLC system. Samples were transported via the autosampler (10uL injection volume) onto a Waters BEH C18 column. Runtime was 15min in PRM mode. Buffer A: 0.1% formic acid in water. Buffer B: 0.1% formic acid in methanol. LC Gradient: 0min: 0% B, 8min: 50% B, 9 min: 98% B, 13min: 98% B. Recalibration of system up to 15 min at 0% B for next injection. |
| Processing Storage Conditions: | Described in summary |
| Extract Storage: | Described in summary |
Combined analysis:
| Analysis ID | AN003829 |
|---|---|
| Analysis type | MS |
| Chromatography type | Reversed phase |
| Chromatography system | Thermo Vanquish |
| Column | Waters Acquity BEH C18 (100 x 2mm,1.7um) |
| MS Type | ESI |
| MS instrument type | Orbitrap |
| MS instrument name | Thermo Orbitrap ID-X tribrid |
| Ion Mode | UNSPECIFIED |
| Units | ug/mL |
Chromatography:
| Chromatography ID: | CH002834 |
| Instrument Name: | Thermo Vanquish |
| Column Name: | Waters Acquity BEH C18 (100 x 2mm,1.7um) |
| Flow Gradient: | 0min: 0% B, 8min: 50% B, 9 min: 98% B, 13min: 98% B. Recalibration of system up to 15 min at 0% B for next injection. |
| Solvent A: | 100% water; 0.1% formic acid |
| Solvent B: | 100% methanol 0.1% formic acid |
| Analytical Time: | 15 min |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS003571 |
| Analysis ID: | AN003829 |
| Instrument Name: | Thermo Orbitrap ID-X tribrid |
| Instrument Type: | Orbitrap |
| MS Type: | ESI |
| MS Comments: | Raw data files were brought into Skyline software. Targeted peak detection was done based on the parent mass. Mass analyzer set to Orbitrap and resolution power set to 120,000 resolution. Then all raw files and unknown samples were imported to Skyline. Calibration curves were generated by Linear regression fit. Targeted precursor MZ and MZ of analytes was used to track and quantification of the metabolite. Peak areas for analytes in samples were used for quantification based in the generated calibration curves. |
| Ion Mode: | UNSPECIFIED |
