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MB Sample ID: SA082452

Local Sample ID:Q7SST_2
Subject ID:SU001253
Subject Type:Cultured cells
Subject Species:Mus musculus
Taxonomy ID:10090

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Subject:

Subject ID:SU001253
Subject Type:Cultured cells
Subject Species:Mus musculus
Taxonomy ID:10090

Factors:

Local Sample IDMB Sample IDFactor Level IDLevel ValueFactor Name
Q7SST_2SA082452FL0125202Time
Q7SST_2SA082452FL012520Q7SST_ControlTreatment

Collection:

Collection ID:CO001247
Collection Summary:STHdhQ111 cells were grown on 10cm dishes in triplicate at a seeding density of 1.06 million cells/well. Compound- or vehicle-treated cells were washed with cold 0.9% NaCl. To each 10cm dish of cells, 660uL LC/MS-grade methanol containing internal standards and 330uL LC/MS-grade water were added. Cells were scraped and transferred to Eppendorf tubes, where 450uL chloroform was added. Samples were vortexed at maximum speed (20,817 rcf) for 10 minutes at 4°C. Each layer was collected separately, avoiding the precipitate at the interface of the two layers, and dried by speedvac.
Sample Type:Cultured cells

Treatment:

Treatment ID:TR001268
Treatment Summary:STHdh cells were incubated in serum-free medium with a compound or vehicle control for 24 hours.

Sample Preparation:

Sampleprep ID:SP001261
Sampleprep Summary:For lipid profiling, cells were resuspended in 50uL 60/35/5 acetonitrile/isopropanol/water (v/v/v) and 5uL was injected for LC/MS analysis. For polar metabolite profiling, cells were resuspended in 100uL water and 2uL was injected for LC/MS analysis.

Combined analysis:

Analysis ID AN001975 AN001976 AN001977 AN001978
Analysis type MS MS MS MS
Chromatography type Reversed phase Reversed phase HILIC HILIC
Chromatography system Thermo Dionex Ultimate 3000 Thermo Dionex Ultimate 3000 Thermo Dionex Ultimate 3000 Thermo Dionex Ultimate 3000
Column Ascentis Express C18 (150 x 2.1mm,2.7um) Ascentis Express C18 (150 x 2.1mm,2.7um) EMD Millipore ZIC-HILIC (100 x 2.1mm,3.5um) EMD Millipore ZIC-HILIC (100 x 2.1mm,3.5um)
MS Type ESI ESI ESI ESI
MS instrument type Orbitrap Orbitrap Orbitrap Orbitrap
MS instrument name Thermo Q Exactive Orbitrap Thermo Q Exactive Orbitrap Thermo Q Exactive Orbitrap Thermo Q Exactive Orbitrap
Ion Mode POSITIVE NEGATIVE POSITIVE NEGATIVE
Units Peak area Peak area Peak Area Peak Area

Chromatography:

Chromatography ID:CH001426
Instrument Name:Thermo Dionex Ultimate 3000
Column Name:Ascentis Express C18 (150 x 2.1mm,2.7um)
Chromatography Type:Reversed phase
  
Chromatography ID:CH001427
Instrument Name:Thermo Dionex Ultimate 3000
Column Name:EMD Millipore ZIC-HILIC (100 x 2.1mm,3.5um)
Chromatography Type:HILIC

MS:

MS ID:MS001829
Analysis ID:AN001975
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:For lipid profiling, cells were resuspended in 50uL 60/35/5 acetonitrile/isopropanol/water (v/v/v) and 5uL was injected for LC/MS analysis. Please see Keckesova et al. and Smulan et al. for a detailed description of the LC/MS analysis (Smulan et al, 2016; Keckesova et al, 2017). Lipid identification and relative quantification was performed using LipidSearch (ThermoFisher Scientific / Mitsui Knowledge Industries). The identified lipids were subjected to quality control filtering and normalization by total signal (Keckesova et al, 2017).
Ion Mode:POSITIVE
  
MS ID:MS001830
Analysis ID:AN001976
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:For lipid profiling, cells were resuspended in 50uL 60/35/5 acetonitrile/isopropanol/water (v/v/v) and 5uL was injected for LC/MS analysis. Please see Keckesova et al. and Smulan et al. for a detailed description of the LC/MS analysis (Smulan et al, 2016; Keckesova et al, 2017). Lipid identification and relative quantification was performed using LipidSearch (ThermoFisher Scientific / Mitsui Knowledge Industries). The identified lipids were subjected to quality control filtering and normalization by total signal (Keckesova et al, 2017).
Ion Mode:NEGATIVE
  
MS ID:MS001831
Analysis ID:AN001977
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:For polar metabolite profiling, cells were resuspended in 100uL water and 2uL was injected for LC/MS analysis. Please see Birsoy et al. and Chen at al. for a detailed description of the LC/MS analysis (Chen et al, 2016; Birsoy et al, 2015). Untargeted analysis was performed using Progenesis CoMet (Nonlinear Dynamics) using the default settings. Features were filtered based on replicate injections and a dilution series of a pooled sample prepared by mixing equal aliquots of the biological samples. Specifically, the filtering criteria were CV < 0.4 across the four replicate injections and R > 0.9 across a four-point dilution series (comprising 0.1X, 0.3X and 1X concentrations, and a double-volume injection). Features that were not lowest according to the Progenesis quantification in the blank water injection samples were discarded.
Ion Mode:POSITIVE
  
MS ID:MS001832
Analysis ID:AN001978
Instrument Name:Thermo Q Exactive Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:For polar metabolite profiling, cells were resuspended in 100uL water and 2uL was injected for LC/MS analysis. Please see Birsoy et al. and Chen at al. for a detailed description of the LC/MS analysis (Chen et al, 2016; Birsoy et al, 2015). Untargeted analysis was performed using Progenesis CoMet (Nonlinear Dynamics) using the default settings. Features were filtered based on replicate injections and a dilution series of a pooled sample prepared by mixing equal aliquots of the biological samples. Specifically, the filtering criteria were CV < 0.4 across the four replicate injections and R > 0.9 across a four-point dilution series (comprising 0.1X, 0.3X and 1X concentrations, and a double-volume injection). Features that were not lowest according to the Progenesis quantification in the blank water injection samples were discarded.
Ion Mode:NEGATIVE
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