A positive/negative ion–switching, targeted mass spectrometry–based metabolomics platform for bodily fluids, cells, and fresh and fixed tissue

By Min Yuan, S. Breitkopf, Xuemei Yang, J. Asara. Nature Protocols, 2012

1,050 citations recorded by the source corpus.

The revival of interest in cancer cell metabolism in recent years has prompted the need for quantitative analytical platforms for studying metabolites from in vivo sources. We implemented a quantitative polar metabolomics profiling platform using selected reaction monitoring with a 5500 QTRAP hybrid triple quadrupole mass spectrometer that covers all major metabolic pathways. The platform uses hydrophilic interaction liquid chromatography with positive/negative ion switching to analyze 258 metabolites (289 Q1/Q3 transitions) from a single 15-min liquid chromatography–mass spectrometry acquisition with a 3-ms dwell time and a 1.55-s duty cycle time. Previous platforms use more than one experiment to profile this number of metabolites from different ionization modes. The platform is compatible with polar metabolites from any biological source, including fresh tissues, cancer cells, bodily fluids and formalin-fixed paraffin-embedded tumor tissue. Relative quantification can be achieved without using internal standards, and integrated peak areas based on total ion current can be used for statistical analyses and pathway analyses across biological sample conditions. The procedure takes ∼12 h from metabolite extraction to peak integration for a data set containing 15 total samples (∼6 h for a single sample).

Subfields: Molecular Biology, Spectroscopy

View Paper: https://www.semanticscholar.org/paper/1f0240d30c15c41389c6dcac5a0755eb43181b0b

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Citations in the corpus

• Metabolomics and isotope tracing (2018, background): “Among these, we find an amide resin to be particularly effective for measuring the core metabolome (Bajad et al., 2006; Yuan et al., 2012).”
• Cognitive Impairment by Antibiotic-Induced Gut Dysbiosis: Analysis of Gut Microbiota-Brain Communication (2016, methodology): “The plasma samples were processed according to Yuan et al. (2012).”
• Vitamin C selectively kills KRAS and BRAF mutant colorectal cancer cells by targeting GAPDH (2015, methodology): “To investigate the mechanism by which vitamin C is selectively toxic to KRAS and BRAF mutant cells, we used LC-MS/MS based metabolomics to profile metabolic changes following vitamin C treatment (17).”
• Consensus guidelines for the use and interpretation of angiogenesis assays (2018, methodology): “For the separation of polar metabolites prior to the MS, two main types of chromatography can be applied: (1) hydrophilic liquid chromatography (HILIC) [177] and (2) ion-pairing chromatography [178].”
• PGC-1α mediates mitochondrial biogenesis and oxidative phosphorylation to promote metastasis (2014, neutral): “Targeted mass spectrometry and metabolomics analyses(18) of 4T1shPGC-1α cells compared to 4T1shScrbl cells revealed lower levels of some of the metabolites levels associated with glycolysis, TCA cycle, amino-sugar metabolism, pyruvate metabolism, phospho-lipid metabolism, pyrimidine and purine metab”