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MB Sample ID: SA151450
Local Sample ID: | 75_B8_D7_RT+ |
Subject ID: | SU001721 |
Subject Type: | Human |
Subject Species: | Homo sapiens |
Taxonomy ID: | 9606 |
Select appropriate tab below to view additional metadata details:
Subject:
Subject ID: | SU001721 |
Subject Type: | Human |
Subject Species: | Homo sapiens |
Taxonomy ID: | 9606 |
Factors:
Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
---|---|---|---|---|
75_B8_D7_RT+ | SA151450 | FL017709 | RT | Treatment |
75_B8_D7_RT+ | SA151450 | FL017709 | 7 | Day |
Collection:
Collection ID: | CO001714 |
Collection Summary: | Whole blood was collected by Canadian Blood Services from donors providing signed informed consent. Buffy coat platelet concentrates suspended in plasma were produced by the Canadian Blood Services Blood4Research Facility in Vancouver, Canada. For each biological replicate, three buffy coat platelet units were pooled and split into three identical platelet units to minimize the effect of donor variability. One unit (RP) was stored at 22°C under constant agitation (Helmer Scientific, PC1200-Pro Platelet Incubator, Noblesville, IN). One unit (CPA) was stored in a walk-in fridge (4°C) in a platelet agitator (Forma platelet agitator, Thermo Electron Corporation. Waltham, MA). The last unit (CP) was stored in walk-in fridge (4°C) without agitation. Platelet units were sampled through a coupling port (Fresenius Kabi, Bad Homburg, Germany) using a sterile syringe in a biosafety cabinet. Platelet units were sampled on days 1, 2, 4, 7, 9, 11, and 14 of storage. This study was approved by both the University of British Columbia Ethics Committee (H17-02708) and the Canadian Blood Services Ethics Board (2018-003). |
Sample Type: | Platelets |
Treatment:
Treatment ID: | TR001734 |
Treatment Summary: | Whole blood was collected by Canadian Blood Services from donors providing signed informed consent. Buffy coat platelet concentrates suspended in plasma were produced by the Canadian Blood Services Blood4Research Facility in Vancouver, Canada. For each biological replicate, three buffy coat platelet units were pooled and split into three identical platelet units to minimize the effect of donor variability. One unit (RP) was stored at 22°C under constant agitation (Helmer Scientific, PC1200-Pro Platelet Incubator, Noblesville, IN). One unit (CPA) was stored in a walk-in fridge (4°C) in a platelet agitator (Forma platelet agitator, Thermo Electron Corporation. Waltham, MA). The last unit (CP) was stored in walk-in fridge (4°C) without agitation. Platelet units were sampled through a coupling port (Fresenius Kabi, Bad Homburg, Germany) using a sterile syringe in a biosafety cabinet. Platelet units were sampled on days 1, 2, 4, 7, 9, 11, and 14 of storage. This study was approved by both the University of British Columbia Ethics Committee (H17-02708) and the Canadian Blood Services Ethics Board (2018-003). |
Sample Preparation:
Sampleprep ID: | SP001727 |
Sampleprep Summary: | A volume of 50μl of platelet concentrates were extracted at a 1:10 dilution in methanol:acetonitrile:water (5:3:2, v/v/v). After vortexing at 4°C for 30 min, extracts were separated from the protein pellet by centrifugation for 10 min at 10,000g at 4°C and stored at −80°C until analysis. For polar metabolites, half of the extracts were dried down via SpeedVac prior to resuspension in ddH2O + 0.1% formic acid, to facilitate ionization and chromatographic resolution of certain polar metabolites (e.g., betaine, kyurenine, etc.). |
Combined analysis:
Analysis ID | AN002689 | AN002690 |
---|---|---|
Analysis type | MS | MS |
Chromatography type | Reversed phase | Reversed phase |
Chromatography system | Thermo Vanquish | Thermo Vanquish |
Column | Phenomenex Kinetex C18 (150 x 2.1mm,2.6um) | Phenomenex Kinetex C18 (150 x 2.1mm,2.6um) |
MS Type | ESI | ESI |
MS instrument type | Orbitrap | Orbitrap |
MS instrument name | Thermo Q Exactive Orbitrap | Thermo Q Exactive Orbitrap |
Ion Mode | NEGATIVE | POSITIVE |
Units | Relative Abundance | Relative Abundance |
Chromatography:
Chromatography ID: | CH001983 |
Chromatography Summary: | UHPLC-MS metabolomics analyses were performed as described,(1, 3) using a Vanquish UHPLC system coupled online to a high-resolution Q Exactive mass spectrometer (Thermo Fisher, Bremen, Germany). Samples were resolved over a Kinetex C18 column (2.1x150 mm, 1.7 µm; Phenomenex, Torrance, CA, USA) at 45°C. A volume of 10 μl of sample extracts was injected into the UHPLC-MS. Each sample was injected and run with two different chromatographic and MS conditions using a 5 minute gradient at 450 µL/minute from 5-95% B (A: 5% acetonitrile, 95%water/1 mM ammonium acetate; B:95%acetonitrile/5% water, 1 mM ammonium acetate) and the MS was operated in negative ion mode. |
Instrument Name: | Thermo Vanquish |
Column Name: | Phenomenex Kinetex C18 (150 x 2.1mm,2.6um) |
Column Temperature: | 45 |
Flow Gradient: | 5 minute gradient from 5-95% B |
Flow Rate: | 450 µL/min |
Solvent A: | 5% acetonitrile/95% water; 1 mM ammonium acetate |
Solvent B: | 95% acetonitrile/5% water; 1 mM ammonium acetate |
Chromatography Type: | Reversed phase |
Chromatography ID: | CH001984 |
Chromatography Summary: | UHPLC-MS metabolomics analyses were performed as described,(1, 3) using a Vanquish UHPLC system coupled online to a high-resolution Q Exactive mass spectrometer (Thermo Fisher, Bremen, Germany). Samples were resolved over a Kinetex C18 column (2.1x150 mm, 1.7 µm; Phenomenex, Torrance, CA, USA) at 45°C. A volume of 10 μl of sample extracts was injected into the UHPLC-MS. Each sample was injected and run with two different chromatographic and MS conditions, using a 5 minute gradient at 450 µL/minute from 5-95% B (A: water/0.1% formic acid; B:acetonitrile/0.1% formic acid) and the MS was operated in positive mode. |
Instrument Name: | Thermo Vanquish |
Column Name: | Phenomenex Kinetex C18 (150 x 2.1mm,2.6um) |
Column Temperature: | 45 |
Flow Gradient: | 5 minute gradient from 5-95% B |
Flow Rate: | 450 µL/min |
Solvent A: | 5% acetonitrile/95% water; 1 mM ammonium acetate |
Solvent B: | 95% acetonitrile/5% water; 1 mM ammonium acetate |
Chromatography Type: | Reversed phase |
MS:
MS ID: | MS002488 |
Analysis ID: | AN002689 |
Instrument Name: | Thermo Q Exactive Orbitrap |
Instrument Type: | Orbitrap |
MS Type: | ESI |
MS Comments: | The UHPLC system was coupled online with a Q Exactive (Thermo, San Jose, CA, USA) scanning in Full MS mode at 70,000 resolution in the 60-900 m/z range, 4 kV spray voltage, 15 sheath gas and 5 auxiliary gas, operated in negative or positive ion mode (separate runs). |
Ion Mode: | NEGATIVE |
MS ID: | MS002489 |
Analysis ID: | AN002690 |
Instrument Name: | Thermo Q Exactive Orbitrap |
Instrument Type: | Orbitrap |
MS Type: | ESI |
MS Comments: | The UHPLC system was coupled online with a Q Exactive (Thermo, San Jose, CA, USA) scanning in Full MS mode at 70,000 resolution in the 60-900 m/z range, 4 kV spray voltage, 15 sheath gas and 5 auxiliary gas, operated in negative or positive ion mode (separate runs). |
Ion Mode: | POSITIVE |