Liquid Chromatography-Mass Spectrometry Metabolic and Lipidomic Sample Preparation Workflow for Suspension-Cultured Mammalian Cells using Jurkat T lymphocyte Cells
- *Corresponding Author:
- Timothy J. Garrett
Department of Pathology
Immunology and Laboratory Medicine
College of Medicine, University of Florida
Gainesville, FL, USA
E-mail: [email protected]
Received date: April 24, 2015; Accepted date: Jun 01, 2015; Published date: June 05, 2015
Citation: Ulmer CZ, Yost RA, Chen J, Mathews CE, Garrett TJ (2015) Liquid Chromatography-Mass Spectrometry Metabolic and Lipidomic Sample Preparation Workflow for Suspension-Cultured Mammalian Cells using Jurkat T lymphocyte Cells. J Proteomics Bioinform 8: 126-132. doi: 10.4172/jpb.1000360
Copyright: © 2015 Ulmer CZ, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Metabolomics is the comprehensive study of metabolism as it pertains to an organism or biological system. Lipidomics, a subset discipline of metabolomics, encompasses the study of cellular lipid functions: including pathways, networks, and interactions. The abundance of metabolites and lipids, along with their contribution to health and disease, makes metabolomics a valuable tool for biomarker research. Disease biomarker identification requires a reproducible, sensitive, and accurate analytical platform. Although transcriptomic and proteomic areas have well-established protocols for sample preparation and data processing, the metabolomics field is still developing comparable standardized conventions. Furthermore, of the few comparative LC-MS metabolomic studies that have been applied to mammalian cell cultures, most are targeted to adherent cell lines. The purpose of this work was to optimize a sample preparation workflow for the cellular metabolomic analysis of suspensioncultured mammalian cells using commercially available Jurkat T lymphocytes as a model system. The current investigation evaluated commonly used sample preparation techniques for reproducibility, accuracy, and applicability to untargeted biomarker discovery. Results show ammoniated cell rinsing solutions to be an effective means to remove extracellular components present in the media without causing ion suppression or affecting the integrity of the cellular membrane. Additionally, a novel workflow was designed to allow for the combined analysis of metabolites and lipids from mammalian suspension cells from a single cell pellet. The Folch lipid extraction protocol was coupled to an 80% MeOH metabolite isolation to ensure high extraction efficiency for phospholipids and triacylglycerides. While the workflow was tailored to cells in suspension it could also be applied to adherent cell lines.