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MB Sample ID: SA229561

Local Sample ID:B1a
Subject ID:SU002415
Subject Type:Human
Subject Species:Homo sapiens
Taxonomy ID:9606
Genotype Strain:N/A
Age Or Age Range:19-75
Weight Or Weight Range:N/A
Height Or Height Range:N/A
Gender:Male and female

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Sample Preparation:

Sampleprep ID:SP002421
Sampleprep Summary:100 μl of saliva samples were thawed, transferred to new tubes, extracted with 200 μl of 80% methanol, and vortexed for 30 seconds. Then the samples were kept at -40 °C for 1 hour, vortexed for 30 s, and centrifuged at 12000 rpm, 4 °C for 15 minutes. Finally, 200 μl of supernatant and 5 μl of DL-o-Chlorophenylalanine (1 mg/ml) were transferred to the vial for LC-MS analysis. Quality control (QC) samples were used to evaluate the methodology. The same amount of extract was obtained from each sample and mixed with QC samples. The QC sample was prepared using the same sample preparation procedure. Instrumental setup Separation was performed by Ultimate 3000LC combined with Q Exactive MS (Thermo) and screened with ESI-MS (targeted MS/MS mode). The LC system is comprised of an ACQUITY UPLC HSS T3 (100 × 2.1mm 1.8 μm) with Ultimate 3000LC. The mobile phase was composed of solvent A (0.05% formic acid-water) and solvent B (acetonitrile) with a gradient elution (0-1.0 min, 95% A; 1.0-12.0 min,95%-5% A; 12.0-13.5 min,5% A; 13.5-13.6 min, 5%-95% A; 13.6-16 min, 95% A). The flow rate of the mobile phase was 0.3 ml/min. The column temperature was maintained at 40 °C, and the sample manager temperature is set at 4 °C. Mass spectrometry parameters in ESI+ and ESI- mode are listed as follows: ESI+: Heater Temp 300 °C; Sheath Gas Flow rate, 45 arb; Aux Gas Flow Rate, 15 arb; Sweep Gas Flow Rate, 1 arb; spray voltage, 3.0 kV; Capillary Temp, 350 °C; S-Lens RF Level, 30%. ESI-: Heater Temp 300 °C, Sheath Gas Flow rate, 45 arb; Aux Gas Flow Rate, 15arb; Sweep Gas Flow Rate, 1 arb; spray voltage, 3.2 kV; Capillary Temp,350 °C; S-Lens RF Level, 60%. Bioinformatic data analysis included multivariate statistical analysis, single variable analysis, cluster analysis, and correlation network of differential metabolites. Statistically significant metabolites (FC >1.5) were integrated with differentially expressed genes obtained from the RNAseq data and the combined data was visualized with Metaboanalyst (https://www.metaboanalyst.ca/) and Cytoscape v3.8 via Metscape v3.1 plugin (https://cytoscape.org/).
Sampleprep Protocol Filename:Sample_preparation.pdf
Processing Storage Conditions:Described in summary
Extract Storage:Described in summary
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