Comprehensive Lipid Profiling of Pancreatic Islets
Pancreatic islets were prepared (SI Appendix) and analyzed using an Agilent 1290 series Ultra-high-performance liquid chromatography system connected to an Agilent 6495 QQQ mass spectrometer after separation on a ZORBAX Eclipse plus C18 column (2.1 × 50 mm, 1.8 µm, 95 Å, Agilent) at 40 °C (SI Appendix). Mass spectrometry analysis was performed in positive ion mode with dynamic, scheduled multiple reaction monitoring (MRM). Mass spectrometry settings, LC-MS gradient, and MRM transitions for each lipid class were adapted from a previously published method (63 (link)). Data analysis was performed on Agilent MassHunter Quantitative analysis software. Relative quantitation was based on one-point calibration with Ceramide d18:1/8:0. The data were further normalized to the total number of islets in each sample.
Zheng X., Ho Q.W., Chua M., Stelmashenko O., Yeo X.Y., Muralidharan S., Torta F., Chew E.G., Lian M.M., Foo J.N., Jung S., Wong S.H., Tan N.S., Tong N., Rutter G.A., Wenk M.R., Silver D.L., Berggren P.O, & Ali Y. (2022). Destabilization of β Cell FIT2 by saturated fatty acids alter lipid droplet numbers and contribute to ER stress and diabetes. Proceedings of the National Academy of Sciences of the United States of America, 119(11), e2113074119.
Ceramide High performance liquid chromatography LipidMass spectrometry Pancreatic islets
Corresponding Organization :
Other organizations :
Sichuan University, Nanyang Technological University, Singapore Eye Research Institute, Singapore General Hospital, Institute of Molecular and Cell Biology, National University of Singapore, Agency for Science, Technology and Research, Genome Institute of Singapore, Imperial College London, Centre Hospitalier de l’Université de Montréal, Karolinska University Hospital, Karolinska Institutet
Separation on a ZORBAX Eclipse plus C18 column (2.1 × 50 mm, 1.8 µm, 95 Å, Agilent) at 40 °C
Mass spectrometry analysis in positive ion mode with dynamic, scheduled multiple reaction monitoring (MRM)
Use of Ceramide d18:1/8:0 as a one-point calibration standard
Normalization of data to the total number of islets in each sample
controls
Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned
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