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MB Sample ID: SA258359
| Local Sample ID: | PTEN WT Rep1 |
| Subject ID: | SU002673 |
| Subject Type: | Mammal |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
| Age Or Age Range: | embryonic fibroblasts |
Select appropriate tab below to view additional metadata details:
Subject:
| Subject ID: | SU002673 |
| Subject Type: | Mammal |
| Subject Species: | Mus musculus |
| Taxonomy ID: | 10090 |
| Age Or Age Range: | embryonic fibroblasts |
Factors:
| Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
|---|---|---|---|---|
| PTEN WT Rep1 | SA258359 | FL033158 | PTEN Wild-Type | Genotype |
Collection:
| Collection ID: | CO002666 |
| Collection Summary: | PTEN WT and KO MEFs were seeded at 80% in 10cm plates 24 hours prior to metabolite extraction/sample prep |
| Sample Type: | Fibroblasts |
Treatment:
| Treatment ID: | TR002685 |
| Treatment Summary: | Cells did not undergo any treatment. They were cultured for 24 hours in 10cm plates with DMEM media containing 10% FBS, 1% Pen/strep, and an additional 1% of glutamine |
Sample Preparation:
| Sampleprep ID: | SP002679 |
| Sampleprep Summary: | Media was fully removed from the plates. Metabolites were extracted using ice cold 80% methanol and dried using a speed vac. The dried metabolites were sent to the Mass Spec Core ran by John Asara at the Beth Israel Deaconess Medical Center at Harvard University for targeted LC-MS/MS. For detailed sample prep information see the attached protocol. |
Combined analysis:
| Analysis ID | AN004237 |
|---|---|
| Analysis type | MS |
| Chromatography type | HILIC |
| Chromatography system | Waters Acquity |
| Column | Waters XBridge Amide (100 x 4.6mm,3.5um) |
| MS Type | APCI |
| MS instrument type | QTRAP |
| MS instrument name | ABI Sciex 5500 QTrap |
| Ion Mode | POSITIVE |
| Units | peak area intensity |
Chromatography:
| Chromatography ID: | CH003144 |
| Instrument Name: | Waters Acquity |
| Column Name: | Waters XBridge Amide (100 x 4.6mm,3.5um) |
| Column Temperature: | 45 |
| Flow Gradient: | 85% buffer B to 30% buffer B from 0 - 3 minutes, then 30% buffer B to 3% buffer B from minute 3 to 12, then 2% buffer B held from minute 12 to 15, followed by 2% buffer B to 85% buffer B from minute 15 to 16, then 85% buffer B held from minute 16-23 in order to re-equilibrate the column. |
| Flow Rate: | 400uL/minute |
| Solvent A: | 95% water/5% acetonitrile; 20 mM ammonium hydroxide; 20 mM ammonium acetate |
| Solvent B: | 100% acetonitrile |
| Chromatography Type: | HILIC |
MS:
| MS ID: | MS003984 |
| Analysis ID: | AN004237 |
| Instrument Name: | ABI Sciex 5500 QTrap |
| Instrument Type: | QTRAP |
| MS Type: | APCI |
| MS Comments: | Selective reaction monitoring, SRM, of endogenous water-soluble metabolites was performed for steady state analysis. The positive ion mode ESI voltage was +4900. Dwell time was set to 3ms per SRM transition and 1.55 seconds was the total cycle time. Retention time 15-20 seconds. MultiQuant v2.1 software was used to in integrate peak areas for each metabolite SRM transition |
| Ion Mode: | POSITIVE |
