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MB Sample ID: SA174189
Local Sample ID: | P H-510 |
Subject ID: | SU001930 |
Subject Type: | Mammal |
Subject Species: | Mesocricetus auratus |
Taxonomy ID: | 10036 |
Select appropriate tab below to view additional metadata details:
Subject:
Subject ID: | SU001930 |
Subject Type: | Mammal |
Subject Species: | Mesocricetus auratus |
Taxonomy ID: | 10036 |
Factors:
Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
---|---|---|---|---|
P H-510 | SA174189 | FL021364 | SARS-CoV-2 | Group |
P H-510 | SA174189 | FL021364 | 14 | Day post infection |
Collection:
Collection ID: | CO001923 |
Collection Summary: | Outbred female LVG golden Syrian hamsters (6-8 weeks of age) were obtained from Charles River Laboratories (Kingston, NY). The hamsters were anesthetized by intraperitoneal injection of a mixture of ketamine and xylazine prior to intranasal inoculation with 0.1 mL of 1e5 plaque-forming units (PFU) of SARS-CoV-2 (WA-1) or H1N1 influenza A virus (A/California/04/2009). On day 2, 4, 6, and 14 after infection, 3-6 anesthetized hamsters per infection group were euthanized by exsanguination followed by intracardiac injection of veterinary euthanasia solution (SleepAway; Fort Dodge). Plasma samples were treated by exposure to germicidal UV-C light. |
Sample Type: | Blood (plasma) |
Treatment:
Treatment ID: | TR001942 |
Treatment Summary: | The hamsters were anesthetized by intraperitoneal injection of a mixture of ketamine and xylazine prior to intranasal inoculation with 0.1 mL of 1e5 plaque-forming units (PFU) of SARS-CoV-2 (WA-1) or H1N1 influenza A virus (A/California/04/2009). |
Sample Preparation:
Sampleprep ID: | SP001936 |
Sampleprep Summary: | Hamster plasma samples were diluted 1:4 with methanol (v/v), vortexed for 30 seconds, and incubated at -20 C for 2 hours. Samples were centrifuged for 10 minutes at 13,500 x g at 4°C and supernatant was transferred to a new centrifuge tube, concentrated, and stored at -80 C until reconstitution. Hamster plasma was thawed on ice. A 50 µL aliquot was transferred onto the solid-phase-extraction (SPE)-system CAPTIVA-EMR Lipid 96-wellplate (Agilent Technologies) before addition of 250 µL of acetonitrile containing 1% formic acid (v/v) and 10 µM internal standard (consisting of uniformly 13C and 15N labeled amino acids from Cambridge Isotope Laboratories, Inc). The samples were mixed for 1 min at 360 rpm on an orbital shaker at room temperature prior to a 10 min incubation period at 4 C. Afterwards, 200 µL 80% acetonitrile in water (v/v) were added to the samples. The samples were mixed on an orbital shaker (360 rpm) for an additional 10 min at room temperature. The samples were then eluted into a 96-deepwell collection plate by centrifugation (10 min, 57 x g, 4 C followed by 2 min, 1000 x g, 4 C). Polar eluates were stored at -80 C until the day of LC/MS analysis. The SPE-plates were then washed twice with 500 µL 80% acetonitrile in water (v/v). Lipids still bound to the SPE-material were then released into a second elution plate, in two elution steps applying 2x 500 µL 1:1 methyl tert-butyl ether:methanol (v/v) onto the SPE cartridge and centrifuging for 2 min at 1000 g and 4 C. The combined eluates were dried under a stream of nitrogen (Biotage SPE Dry Evaporation System) at room temperature and reconstituted with 100 µL 1:1 2-propanol:methanol (v/v) prior to LC/MS analysis. |
Combined analysis:
Analysis ID | AN003001 | AN003002 | AN003003 | AN003004 |
---|---|---|---|---|
Analysis type | MS | MS | MS | MS |
Chromatography type | HILIC | HILIC | Reversed phase | Reversed phase |
Chromatography system | Agilent 1290 Infinity II | Agilent 1290 Infinity II | Agilent 1290 Infinity II | Agilent 1290 Infinity II |
Column | SeQuant ZIC-pHILIC (100 x 2.1mm,5um) including a ZIC-pHILIC guard (2.1 mm x 20 mm,5um) | SeQuant ZIC-pHILIC (100 x 2.1mm,5um) including a ZIC-pHILIC guard (2.1 mm x 20 mm,5um) | Waters ACQUITY UPLC HSS T3 (150 x 2.1mm,1.8um) including an Acquity UPLC® HSS T3 VanGuard Pre-Column (2.1 x 5mm,1.8 µm) | Waters ACQUITY UPLC HSS T3 (150 x 2.1mm,1.8um) including an Acquity UPLC® HSS T3 VanGuard Pre-Column (2.1 x 5mm,1.8 µm) |
MS Type | ESI | ESI | ESI | ESI |
MS instrument type | QTOF | QTOF | QTOF | QTOF |
MS instrument name | Agilent 6540 QTOF | Agilent 6540 QTOF | Agilent 6545 QTOF | Agilent 6545 QTOF |
Ion Mode | POSITIVE | NEGATIVE | POSITIVE | NEGATIVE |
Units | Relative Intensity | Relative Intensity | Relative Intensity | Relative Intensity |
Chromatography:
Chromatography ID: | CH002226 |
Chromatography Summary: | An aliquot of 2 µL of polar metabolite extract was subjected to LC/MS analysis by using an Agilent 1290 Infinity II liquid-chromatography (LC) system coupled to an Agilent 6540 Quadrupole-Time-of-Flight (Q-TOF) mass spectrometer with a dual Agilent Jet Stream electrospray ionization source. Polar metabolites were separated on a SeQuant® ZIC®-pHILIC column (100 x 2.1 mm, 5 µm, polymer, Merck-Millipore) including a ZIC®-pHILIC guard column (2.1 mm x 20 mm, 5 µm). The column compartment temperature was maintained at 40 C and the flow rate was set to 250 µLmin-1. The mobile phases consisted of A: 95% water, 5% acetonitrile, 20 mM ammonium bicarbonate, 0.1% ammonium hydroxide solution (25% ammonia in water), 2.5 µM medronic acid, and B: 95% acetonitrile, 5% water, 2.5 µM medronic acid. The following linear gradient was applied: 0 to 1 min, 90% B; 12 min, 35% B; 12.5 to 14.5 min, 25% B; 15 min, 90% B followed by a re-equilibration phase of 4 min at 400 µLmin-1 and 2 min at 250 µLmin-1. |
Instrument Name: | Agilent 1290 Infinity II |
Column Name: | SeQuant ZIC-pHILIC (100 x 2.1mm,5um) including a ZIC-pHILIC guard (2.1 mm x 20 mm,5um) |
Column Temperature: | 40 |
Flow Gradient: | 0 to 1 min, 90% B; 12 min, 35% B; 12.5 to 14.5 min, 25% B; 15 min, 90% B followed by a re-equilibration phase of 4 min |
Flow Rate: | 250ul/min |
Solvent A: | 95% water/5% acetonitrile; 20 mM ammonium bicarbonate; 0.1% ammonium hydroxide; 2.5 µM medronic acid |
Solvent B: | 95% acetonitrile/5% water; 2.5 µM medronic acid |
Chromatography Type: | HILIC |
Chromatography ID: | CH002227 |
Chromatography Summary: | An aliquot of 2 µL of lipid extract was subjected to LC/MS analysis by using an Agilent 1290 Infinity II LC-system coupled to an Agilent 6545 Q-TOF mass spectrometer with a dual Agilent Jet Stream electrospray ionization source. Lipids were separated on an Acquity UPLC® HSS T3 column (2.1 x 150 mm, 1.8 µm) including an Acquity UPLC® HSS T3 VanGuard Pre-Column (2.1 x 5mm, 1.8 µm) at a temperature of 60 C and a flow rate of 250 µLmin-1. The mobile phases consisted of A: 60% acetonitrile, 40% water, 0.1% formic acid, 10 mM ammonium formate, 2.5 µM medronic acid, and B: 90% 2-propanol, 10% acetonitrile, 0.1% formic acid, 10 mM ammonium formate (dissolved in 1 mL water). The following linear gradient was used: 0-2 min, 30% B; 17 min, 75% B; 20 min, 85%; 23-26 min, 100% B; 26, 30% B followed by a re-equilibration phase of 5 min. |
Instrument Name: | Agilent 1290 Infinity II |
Column Name: | Waters ACQUITY UPLC HSS T3 (150 x 2.1mm,1.8um) including an Acquity UPLC® HSS T3 VanGuard Pre-Column (2.1 x 5mm,1.8 µm) |
Column Temperature: | 60 |
Flow Gradient: | The following linear gradient was used: 0-2 min, 30% B; 17 min, 75% B; 20 min, 85%; 23-26 min, 100% B; 26, 30% B followed by a re-equilibration phase of 5 min. |
Flow Rate: | 0.25ml/min |
Solvent A: | 60% acetonitrile/40% water; 0.1% formic acid; 10mM ammonium formate; 2.5uM medronic acid |
Solvent B: | 90% isopropanol/10% acetonitrile; 0.1% formic acid; 10 mM ammonium formate |
Chromatography Type: | Reversed phase |
MS:
MS ID: | MS002790 |
Analysis ID: | AN003001 |
Instrument Name: | Agilent 6540 QTOF |
Instrument Type: | QTOF |
MS Type: | ESI |
MS Comments: | gas temperature 200 C, drying gas flow 10 L*min-1, nebulizer pressure 44 psi, sheath gas temperature 300 C, sheath gas flow 12 L*min-1, VCap 3000 V, nozzle voltage 2000 V, Fragmentor 100 V, Skimmer 65 V, Oct 1 RF Vpp 750 V, and m/z range 50-1700. Data were acquired under continuous reference mass correction at m/z 121.0509 and 922.0890 for positive ion mode and m/z 119.0363 and 966.0007 for negative ion mode. Polar metabolite identifications were supported by matching the retention time, accurate mass, and MS/MS fragmentation data to our in-house retention time and MS/MS library created from authentic reference standards (Mass Spectrometry Metabolite Library supplied by IROA Technologies, Millipore Sigma, St. Louis, MO, USA) and online MS/MS libraries (Human Metabolome Database (HMDB, https://hmdb.ca), Mass Bank of North America (MoNA, https://mona.fiehnlab.ucdavis.edu/), and mzCloud (https://mzcloud.org). Lipid iterative MS/MS data were annotated with the Agilent Lipid Annotator software. All data files were then analyzed in Skyline (Version 20.1.0.155) to obtain peak areas. |
Ion Mode: | POSITIVE |
MS ID: | MS002791 |
Analysis ID: | AN003002 |
Instrument Name: | Agilent 6540 QTOF |
Instrument Type: | QTOF |
MS Type: | ESI |
MS Comments: | gas temperature 200 C, drying gas flow 10 L*min-1, nebulizer pressure 44 psi, sheath gas temperature 300 C, sheath gas flow 12 L*min-1, VCap 3000 V, nozzle voltage 2000 V, Fragmentor 100 V, Skimmer 65 V, Oct 1 RF Vpp 750 V, and m/z range 50-1700. Data were acquired under continuous reference mass correction at m/z 121.0509 and 922.0890 for positive ion mode and m/z 119.0363 and 966.0007 for negative ion mode. Polar metabolite identifications were supported by matching the retention time, accurate mass, and MS/MS fragmentation data to our in-house retention time and MS/MS library created from authentic reference standards (Mass Spectrometry Metabolite Library supplied by IROA Technologies, Millipore Sigma, St. Louis, MO, USA) and online MS/MS libraries (Human Metabolome Database (HMDB, https://hmdb.ca), Mass Bank of North America (MoNA, https://mona.fiehnlab.ucdavis.edu/), and mzCloud (https://mzcloud.org). Lipid iterative MS/MS data were annotated with the Agilent Lipid Annotator software. All data files were then analyzed in Skyline (Version 20.1.0.155) to obtain peak areas. |
Ion Mode: | NEGATIVE |
MS ID: | MS002792 |
Analysis ID: | AN003003 |
Instrument Name: | Agilent 6545 QTOF |
Instrument Type: | QTOF |
MS Type: | ESI |
MS Comments: | gas temperature 250 C, drying gas flow 11 L*min-1, nebulizer pressure 35 psi, sheath gas temperature 300 C, sheath gas flow 12 L*min-1, VCap 3000 V, nozzle voltage 500 V, Fragmentor 160 V, Skimmer 65 V, Oct 1 RF Vpp 750 V, and m/z range 50-1700. Data were acquired under continuous reference mass correction at m/z 121.0509 and 922.0890 in positive ion mode and m/z 119.0363 and 966.0007 in negative ion mode. Polar metabolite identifications were supported by matching the retention time, accurate mass, and MS/MS fragmentation data to our in-house retention time and MS/MS library created from authentic reference standards (Mass Spectrometry Metabolite Library supplied by IROA Technologies, Millipore Sigma, St. Louis, MO, USA) and online MS/MS libraries (Human Metabolome Database (HMDB, https://hmdb.ca), Mass Bank of North America (MoNA, https://mona.fiehnlab.ucdavis.edu/), and mzCloud (https://mzcloud.org). Lipid iterative MS/MS data were annotated with the Agilent Lipid Annotator software. All data files were then analyzed in Skyline (Version 20.1.0.155) to obtain peak areas. |
Ion Mode: | POSITIVE |
MS ID: | MS002793 |
Analysis ID: | AN003004 |
Instrument Name: | Agilent 6545 QTOF |
Instrument Type: | QTOF |
MS Type: | ESI |
MS Comments: | gas temperature 250 C, drying gas flow 11 L*min-1, nebulizer pressure 35 psi, sheath gas temperature 300 C, sheath gas flow 12 L*min-1, VCap 3000 V, nozzle voltage 500 V, Fragmentor 160 V, Skimmer 65 V, Oct 1 RF Vpp 750 V, and m/z range 50-1700. Data were acquired under continuous reference mass correction at m/z 121.0509 and 922.0890 in positive ion mode and m/z 119.0363 and 966.0007 in negative ion mode. Polar metabolite identifications were supported by matching the retention time, accurate mass, and MS/MS fragmentation data to our in-house retention time and MS/MS library created from authentic reference standards (Mass Spectrometry Metabolite Library supplied by IROA Technologies, Millipore Sigma, St. Louis, MO, USA) and online MS/MS libraries (Human Metabolome Database (HMDB, https://hmdb.ca), Mass Bank of North America (MoNA, https://mona.fiehnlab.ucdavis.edu/), and mzCloud (https://mzcloud.org). Lipid iterative MS/MS data were annotated with the Agilent Lipid Annotator software. All data files were then analyzed in Skyline (Version 20.1.0.155) to obtain peak areas. |
Ion Mode: | NEGATIVE |