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MB Sample ID: SA316328
Local Sample ID: | MEL_100nM_Timecourse_Day2_HILIC_3 |
Subject ID: | SU003025 |
Subject Type: | Cultured cells |
Subject Species: | Mus musculus |
Taxonomy ID: | 10090 |
Cell Strain Details: | MEL |
Select appropriate tab below to view additional metadata details:
Subject:
Subject ID: | SU003025 |
Subject Type: | Cultured cells |
Subject Species: | Mus musculus |
Taxonomy ID: | 10090 |
Cell Strain Details: | MEL |
Factors:
Local Sample ID | MB Sample ID | Factor Level ID | Level Value | Factor Name |
---|---|---|---|---|
MEL_100nM_Timecourse_Day2_HILIC_3 | SA316328 | FL037612 | Day2 | 100nM_Folic_Acid_Growth |
Collection:
Collection ID: | CO003018 |
Collection Summary: | One million cells from culture were collected via centrifugation for 20 seconds at 18,000xG, washed with 0.9% NaCl, and collected via centrifugation for 20 seconds at 18,000xG |
Sample Type: | Cultured cells |
Treatment:
Treatment ID: | TR003034 |
Treatment Summary: | MEL cells were cultured for up to 8 days in 100 nM folic acid containing RPMI media. The treatments were setup as a reverse time-course experiment and harvested on the same day. Day 0 samples were cultured for 8 days in 2,000 nM folic acid. |
Sample Preparation:
Sampleprep ID: | SP003031 |
Sampleprep Summary: | Pellet was resuspended in extraction buffer (80% Methanol, 25 mM Ammonium Acetate and 2.5 mM Na-Ascorbate prepared in LC-MS water, supplemented with isotopically-labelled amino acid standards [Cambridge Isotope Laboratories, MSK-A2-1.2], aminopterin, and reduced glutathione standard [Cambridge Isotope Laboratories, CNLM-6245-10]). Samples were vortexed for 10 sec, then centrifuged for 10 minutes at 18,000 g to pellet cell debris. The supernatant was divided into two tubes and dried on ice using a liquid nitrogen dryer. One tube of dried sample was used for polar metabolite detection. Dried samples were resuspended in 25 μL water |
Combined analysis:
Analysis ID | AN004782 |
---|---|
Analysis type | MS |
Chromatography type | HILIC |
Chromatography system | Thermo Vanquish |
Column | EMD Millipore ZIC-HILIC (100 x 2.1 mm,3.5 um) |
MS Type | ESI |
MS instrument type | Orbitrap |
MS instrument name | Thermo Q Exactive Orbitrap |
Ion Mode | UNSPECIFIED |
Units | peak area |
Chromatography:
Chromatography ID: | CH003613 |
Chromatography Summary: | Sample was injected into a ZIC-pHILIC 150 x 2.1 mm (5 μm particle size) column (EMD Millipore). operated on a Vanquish™ Flex UHPLC Systems (Thermo Fisher Scientific, San Jose, CA, USA). Chromatographic separation was achieved using the following conditions: buffer A was acetonitrile; buffer B was 20 mM ammonium carbonate, 0.1% ammonium hydroxide in water; resulting pH is around 9 without pH adjustment. Gradient conditions we used were: linear gradient from 20% to 80% B; 20–20.5 min: from 80% to 20% B; 20.5–28 min: hold at 20% B at 150 mL/min flow rate. The column oven and autosampler tray were held at 25 °C and 4 °C, respectively. |
Instrument Name: | Thermo Vanquish |
Column Name: | EMD Millipore ZIC-HILIC (100 x 2.1 mm,3.5 um) |
Column Temperature: | 25 |
Flow Gradient: | linear gradient from 20% to 80% B; 20–20.5 min: from 80% to 20% B; 20.5–28 min: hold at 20% B at 150 mL/min flow rate |
Flow Rate: | 150 mL/min |
Solvent A: | 100% acetonitrile |
Solvent B: | 100% water; 20 mM ammonium carbonate; 0.1% ammonium hydroxide |
Chromatography Type: | HILIC |
MS:
MS ID: | MS004528 |
Analysis ID: | AN004782 |
Instrument Name: | Thermo Q Exactive Orbitrap |
Instrument Type: | Orbitrap |
MS Type: | ESI |
MS Comments: | Polar metabolites were relatively quantified while referencing an in-house library of chemical standards and using TraceFinder 4.1 (Thermo Fisher Scientific, Waltham, MA, USA), with a 5 ppm mass tolerance. |
Ion Mode: | UNSPECIFIED |