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MB Sample ID: SA248053
Local Sample ID: | PMN_63 |
Subject ID: | SU002567 |
Subject Type: | Human |
Subject Species: | Homo sapiens |
Taxonomy ID: | 9606 |
Select appropriate tab below to view additional metadata details:
Subject:
Subject ID: | SU002567 |
Subject Type: | Human |
Subject Species: | Homo sapiens |
Taxonomy ID: | 9606 |
Factors:
Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
---|---|---|---|---|
PMN_63 | SA248053 | FL031177 | Severe | Experimental Factors |
Collection:
Collection ID: | CO002560 |
Collection Summary: | COVID-19 patient samples were obtained through the University of Texas Southwestern Medical Center biorepository. Sample collection from COVID-19 patients or healthy controls was performed according to procedures approved by the Institutional Review Board of the University of Texas Southwestern Medical Center (STU-2020-0375 for SARS-CoV2 Biorepository and STU 012014-040 to J. Moreland) and the UT Southwestern Biorepository. Neutrophils were isolated using Ficoll-paque density gradient centrifugation, dextran sedimentation and erythrocyte hypotonic lysis following standard procedures as previously described (Hook et al., 2021). The number of neutrophils in each sample was counted using a microscope before freezing. Samples were stored at -800 C until metabolite extraction. |
Sample Type: | Neutrophils |
Treatment:
Treatment ID: | TR002579 |
Treatment Summary: | No treatment were done |
Sample Preparation:
Sampleprep ID: | SP002573 |
Sampleprep Summary: | Metabolites from pellets containing 2 million neutrophils were extracted with 100 μl of chilled solvent of 40:40:20 methanol:acetonitrile:water (v:v:v) per million cells. The cell pellet was resuspended in the extraction solvent, vortexed for 30’’, and freeze-thawed 3 times. Then the samples were centrifuged at 21,000 g for 15 minutes at 4 °C and the supernatant transferred to LC-MS autosampler vials. |
Combined analysis:
Analysis ID | AN004046 |
---|---|
Analysis type | MS |
Chromatography type | HILIC |
Chromatography system | Thermo Scientific QExactive HF-X hybrid quadrupole orbitrap high-resolution mass spectrometer (HRMS) |
Column | Millipore ZIC-pHILIC column (5 mm, 2.1×150 mm) |
MS Type | ESI |
MS instrument type | Orbitrap |
MS instrument name | Thermo Q Exactive HF-X Orbitrap |
Ion Mode | UNSPECIFIED |
Units | MS reading |
Chromatography:
Chromatography ID: | CH002993 |
Chromatography Summary: | Liquid chromatography was performed using a Millipore ZIC-pHILIC column (5 mm, 2.1×150 mm) with a binary solvent system of 10 mM ammonium acetate in water, pH 9.8 (solvent A) and acetonitrile (solvent B) with a constant flow rate of 0.25 mL/min. The column was equilibrated with 90% solvent B. The liquid chromatography gradient was: 0–15 minutes linear ramp from 90% B to 30% B; 15–18 minutes isocratic flow of 30% B; 18–19 minutes linear ramp from 30% B to 90% B; 19–27 column regeneration with isocratic flow of 90% B. |
Instrument Name: | Thermo Scientific QExactive HF-X hybrid quadrupole orbitrap high-resolution mass spectrometer (HRMS) |
Column Name: | Millipore ZIC-pHILIC column (5 mm, 2.1×150 mm) |
Column Temperature: | 40 |
Flow Gradient: | 0–15 minutes linear ramp from 90% B to 30% B; 15–18 minutes isocratic flow of 30% B; 18–19 minutes linear ramp from 30% B to 90% B; 19–27 column regeneration with isocratic flow of 90% B. |
Flow Rate: | 0.25 mL/min |
Solvent A: | 100% water; 10 mM ammonium acetate, pH 9.8 |
Solvent B: | 100% acetonitrile |
Chromatography Type: | HILIC |
MS:
MS ID: | MS003793 |
Analysis ID: | AN004046 |
Instrument Name: | Thermo Q Exactive HF-X Orbitrap |
Instrument Type: | Orbitrap |
MS Type: | ESI |
MS Comments: | Polarity switching was used to detect both positive and negative ions. A pooled quality control (QC) sample was used through the LC-MS run interspersed with the neutrophil samples to monitor the quality and consistency of detection. All samples were randomized to avoid artifactual changes caused by systematic drift. For data acquired with the QExactive, chromatogram peak areas were integrated using Tracefinder (Thermo Scientific) for targeted analysis using an in-house database. |
Ion Mode: | UNSPECIFIED |