Untargeted lipidomics of CT26 cells
Description
Untargeted lipidomics of 37°C-CT26 and 45°C-CT26 cells
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Steps to reproduce
Freshly collected 37°C-CT26 and 45°C-CT26 cells (1 × 107 cells/per sample, n = 6) were resuspended in 1 mL of PBS, snapped frozen and saved at -80°C until processing. To the sample 1 mL of lipid extract was added, followed by votexing and centrifugation at 12000 rpm for 10 min at 4°C. The organic fraction was transferred to a clean vial and dried under N2, followed by reconstitution in 100 mL acetonitrile (ACN)/isopropanol (IPA). For Untargeted Lipidomics LC-MS, reconstituted extracts were injected into a Ultra Performance Liquid Chromatography (UPLC) (Shim-pack UFLC SHIMADZU CBM30A) coupled to a Tandem mass spectrometry (QTRAP® 6500+). Lipids were separated using a Thermo C30 column (2.1 × 100 mm) with the following solvents A: 5 mmol/L ammonium formate with 0.04% acetic acid in 60:40 ACN: water and B: 5 mmol/L ammonium formate with 0.04% acetic acid in 10:90 ACN: IPA at a flow rate of 0.35 mL/min. The gradient program was 0 min A/B (80:20 V/V), 3 min 50:50 V/V, 5 min 35:65 V/V, 9 min 25:75 V/V, and 15.5 min 10:90 V/V. The mass spectrometer was equipped with an ESI source. Statistically significant was defined according to the criteria of | log2(fold change) | > 1.