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MB Sample ID: SA237309
Local Sample ID: | serumPBQC_3 |
Subject ID: | SU002459 |
Subject Type: | Mammal |
Subject Species: | Mus musculus |
Taxonomy ID: | 10090 |
Select appropriate tab below to view additional metadata details:
Subject:
Subject ID: | SU002459 |
Subject Type: | Mammal |
Subject Species: | Mus musculus |
Taxonomy ID: | 10090 |
Factors:
Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
---|---|---|---|---|
serumPBQC_3 | SA237309 | FL029382 | Pooled Biological Quality Control | Treatment |
Collection:
Collection ID: | CO002452 |
Collection Summary: | Groups of male mice were treated with 4 x daily injections of Colony Stimulating Factor (1), followed by normal feeding or a 24 h fast from day 6 to day 7. Serum was obtained from blood obtained from cardiac puncture under isofluorane anaesthesia. |
Sample Type: | serum |
Treatment:
Treatment ID: | TR002471 |
Treatment Summary: | Groups of male mice were treated with 4 x daily injections of Colony Stimulating Factor (1), followed by normal feeding or a 24 h fast from day 6 to day 7. |
Sample Preparation:
Sampleprep ID: | SP002465 |
Sampleprep Summary: | Metabolites were extracted from 50 µl serum using 150 µl methanol and 50 µl chloroform, including 13C-sorbitol and 13C,15N-valine internal standards, and 30 µl of clarified sera was prepared for GC-MS analysis. Liver metabolites were extracted from 20-40 mg liver tissue (sampled from the same region of the left lateral lobe) in 600 µl methanol:MilliQ water (3:1), including 13C-sorbitol and 13C,15N-valine internal standards. The tissue was cryomilled at 4°C. 110 µl chloroform was added to 440 µg homogenate which was transferred to a new tube and the sample thermomixed for 20 minutes then centrifuged at 13,000 rpm for 15 minutes. 30 µl supernatant was dried in inserts for GC-MS analysis. Dried samples for targeted analysis were derivatised online using the Shimadzu AOC6000 autosampler robot. Derivatisation was achieved by the addition of 25 µL methoxyamine hydrochloride (30 mg/mL in pyridine, Merck) followed by shaking at 37°C for 2h. Samples were then derivatised with 25 µL of N,O-bis (trimethylsilyl)trifluoroacetamide with trimethylchlorosilane (BSTFA with 1% TMCS, Thermo Scientific) for 1h at 37°C. The sample was allowed to equilibrate at room temperature for 1 h before 1 µL was injected onto the GC column using a hot needle technique. Split (1:10) injections were performed for each sample. |
Combined analysis:
Analysis ID | AN003865 |
---|---|
Analysis type | MS |
Chromatography type | None (Direct infusion) |
Chromatography system | Shimadzu TQ8050NX |
Column | Agilent DB5-MS (30m) |
MS Type | ESI |
MS instrument type | Triple quadrupole |
MS instrument name | new |
Ion Mode | UNSPECIFIED |
Units | area |
Chromatography:
Chromatography ID: | CH002863 |
Instrument Name: | Shimadzu TQ8050NX |
Column Name: | Agilent DB5-MS (30m) |
Chromatography Type: | None (Direct infusion) |
MS:
MS ID: | MS003606 |
Analysis ID: | AN003865 |
Instrument Name: | new |
Instrument Type: | Triple quadrupole |
MS Type: | ESI |
MS Comments: | The GC-MS system used comprised of an AOC6000 autosampler, a 2030 Shimadzu gas chromatograph and a TQ8050NX triple quadrupole mass spectrometer (Shimadzu, Japan). The mass spectrometer was tuned according to the manufacturer’s recommendations using tris-(perfluorobutyl)-amine (CF43). GC-MS was performed on a 30m Agilent DB-5 column with 0.25mm internal diameter column and 1µm film thickness. The injection temperature (inlet) was set at 280°C, the MS transfer line at 280°C and the ion source adjusted to 200°C. Helium was used as the carrier gas at a flow rate of 1 mL/min and argon gas was used in the collision cell to generate the MRM product ion. The analysis of TMS samples was performed under the following oven temperature program; 100°C start temperature, hold for 4 minutes, followed by a 10°C min-1 oven temperature ramp to 320°C with a following final hold for 11 minutes. Approximately 520 targets were collected using the Shimadzu Smart Metabolite Database, where each target comprised a quantifier MRM along with a qualifier MRM, which covers approximately 350 endogenous metabolites and multiple stable isotopically labelled internal standards. Resultant data was processed using Shimadzu LabSolutions Insight software, where peak integrations were visually validated and manually corrected where required. |
Ion Mode: | UNSPECIFIED |