Summary of Study ST001676
This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR001078. The data can be accessed directly via it's Project DOI: 10.21228/M8C111 This work is supported by NIH grant, U2C- DK119886.
See: https://www.metabolomicsworkbench.org/about/howtocite.php
This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.
| Study ID | ST001676 |
| Study Title | Lipidomic analysis of CD4+ T-cell subsets (Th1,Th2,Th17 and iTreg cells) (part I) |
| Study Type | MS, untargeted cell-based lipidomics |
| Study Summary | Part 1/5: It includes lipidomic analysis of CD4+ T-cell subsets(Th1,Th2,Th17 and iTreg cells)and their paired controls(Th0 cells). |
| Institute | University of Turku |
| Department | Systems Medicine, Turku Bioscience |
| Laboratory | Systems Medicine |
| Last Name | Sen |
| First Name | Partho |
| Address | Tykistökatu 6B, BioCity, 5th Floor, Turku, Southwest, 20521, Finland |
| partho.sen@utu.fi | |
| Phone | 0469608145 |
| Submit Date | 2021-01-28 |
| Raw Data Available | Yes |
| Raw Data File Type(s) | mzML |
| Analysis Type Detail | LC-MS |
| Release Date | 2021-11-02 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Combined analysis:
| Analysis ID | AN002734 |
|---|---|
| Analysis type | MS |
| Chromatography type | Reversed phase |
| Chromatography system | Waters Acquity UHPLC UNSPSC 41115709 |
| Column | Waters BEH C18 (00mm x 2.1mm,1.7um ) |
| MS Type | ESI |
| MS instrument type | QTOF |
| MS instrument name | Agilent 6540 QTOF |
| Ion Mode | POSITIVE |
| Units | Intensities |
MS:
| MS ID: | MS002531 |
| Analysis ID: | AN002734 |
| Instrument Name: | Agilent 6540 QTOF |
| Instrument Type: | QTOF |
| MS Type: | ESI |
| MS Comments: | The UHPLC-QTOFMS analyses were done with some modifications on two separate instruments. The initial lipidomic results were acquired on a UHPLC-QTOFMS system from Agilent Technologies (Santa Clara, CA, USA) combining a 1290 Infinity LC system and 6545 quadrupole time of flight mass spectrometer (QTOFMS), interfaced with a dual jet stream electrospray (dual ESI) ion source. MassHunter B.06.01 software (Agilent Technologies, Santa Clara, CA, USA) was used for all data acquisition. The SM results for UGCG-silenced Th17 cells data was acquired on a UHPLC-QTOF system from Bruker (Bruker, Billerica, MA, USA) combining an Elute UHPLC binary pump and an Impact II system QTOF system. The samples for this experiments were the same extracts that the ceramide (Cer) data was acquired from and had SM(18:1/17:0) spiked in prior to acquisition. The data was acquired using the Hystar suite of software. MZmine 2 was used for all the untargeted data processing. MS data were processed using the open source software MZmine 2.53. The following data processing steps were applied to the raw MS data: (1) Crop filtering with a m/z range of 350 – 1200 m/z and a retention time (RT) range of 2 to 15 minutes; (2) Mass detection with a noise level of 900; (3) Chromatogram builder with a min time span of 0.08 minutes, minimum height of 900 and m/z tolerance of 0.006 m/z or 10.0 ppm; (4) Chromatogram deconvolution using the local minimum search algorithm with a 70% chromatographic threshold, 0.05 min minimum RT range, 5% minimum relative height, 1200 minimum absolute height, a minimum ration of peak top/edge of 1.2 and a peak duration range of 0.08 - 1.01 minutes; (5) Isotopic peak grouper with a m/z tolerance of 5.0 ppm, RT tolerance of 0.05 minute, maximum charge of 2 and with the most intense isotope set as the representative isotope; (6) Join aligner with m/z tolerance of 0.009 or 10.0 ppm and a weight of 2, RT tolerance of 0.1 minute and a weight of 1 and with no requirement of charge state or ID and no comparison of isotope pattern; (7) Peak list row filter with a minimum of 7 peaks in a row (10% of the samples); (8) Gap filling using the same RT and m/z range gap filler algorithm with an m/z tolerance of 0.009 m/z or 11.0 ppm; (9) Identification of lipids using a custom database (based on UHPLC-MS/MS data using the same lipidomics protocol, with RT data and MS and MS/MS) search with an m/z tolerance of 0.009 m/z or 10.0 ppm and a RT tolerance of 0.2 min. In general, lipids were identified at the total number of carbons and double bonds in the structure as there was insufficient evidence to assign the specific acyl chains. Where the acyl chains are identified these have been confirmed with MS/MS level experiments and/or authentic standards. (10) Normalization using internal standards (PE (17:0/17:0), SM (d18:1/17:0), Cer (d18:1/17:0), LPC (17:0), TG (17:0/17:0/17:0) and PC (16:0/d30/18:1)) for identified lipids and closest internal standard (based on RT) for the unknown lipids, followed by calculation of the concentrations based on lipid-class calibration curves. Identification of lipids was done using an in-house spectral library with MS (and retention time), MS/MS information, and by searching the LIPID MAPS spectral database (http://www.lipidmaps.org). MS/MS data were acquired in both negative and positive ion modes in order to maximize identification coverage. Additionally, some lipids were verified by injection of commercial standards. The identification was carried out in pooled cell extracts. The peak area obtained for each lipid was normalized with lipid-class specific internal standards and with total content of protein. A (semi) quantitation was performed using lipid-class specific calibration curves. Pooled cell extracts were used for quality control, in addition to in-house plasma. The raw variation of the peak areas of internal standards in the samples was on average 15.3% and the RSD of retention times of identified lipids across all samples was on average 0.28%. The RSD of the concentrations of the identified lipids in QC samples and pooled extracts was on average 17.7%. |
| Ion Mode: | POSITIVE |
