Flux will be measured using the metabolomics platforms based on absolute quantification method (isotope ratio based MS technique) by LC-MS, using heavy-isotope labelled precursors of the metabolites of interest. For example, 15N labelled cysteine, glycine and glutamate will be used to determine rates of synthesis of glutathione. 15N-labelled methionine to measure S-adenosyl methionine (and its decarboxylated form, as well as methionine cycle intermediates). 15N labelled arginine is used as precursor for ornithine, putrescine, and spermidine and trypanothione synthesis. 13C labelled glucose will be used to measure production of glucose 6-phosphate, 6-phosphogluconate and pentose-phosphate. Experiments will involve adding these precursors to medium and then following their distribution into the different metabolites under normal physiological conditions and comparing that distribution over time with cells perturbed with the different stress inducing agents and also in selected RNAi and gene knockout cells defective in various enzymes.
SEEK ID: https://fairdomhub.org/studies/104
Metabolite profiling, quantification and flux quantification
Projects: SilicoTryp
Study position:
Experimentalists: Dong-Hyun Kim
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Created: 10th Dec 2012 at 16:13
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Projects: SilicoTryp
Institutions: University of Glasgow
Expertise: Metabolomics
Tools: Metabolomics, Cell culture, GC and LC/MS analysis of metabolites
SysMO is a European transnational funding and research initiative on "Systems Biology of Microorganisms".
The goal pursued by SysMO was to record and describe the dynamic molecular processes going on in unicellular microorganisms in a comprehensive way and to present these processes in the form of computerized mathematical models.
Systems biology will raise biomedical and biotechnological research to a new quality level and contribute markedly to progress in understanding. Pooling European research ...
Projects: BaCell-SysMO, COSMIC, SUMO, KOSMOBAC, SysMO-LAB, PSYSMO, SCaRAB, MOSES, TRANSLUCENT, STREAM, SulfoSys, SysMO DB, SysMO Funders, SilicoTryp, Noisy-Strep
Web page: http://sysmo.net/
The SilicoTryp project aims at the creation of a “Silicon Trypanosome”, a comprehensive, experiment-based, multi-scale mathematical model of trypanosome physiology. Trypanosomes are blood-stream parasites transmitted by tsetse flies; they cause African sleeping sickness in humans and livestock. Currently available drugs have severe side effects, and the parasites are rapidly developing resistance. In this project, we collect a wide range of new experimental data on the parasite in its various ...
Programme: SysMO
Public web page: http://silicotryp.ibls.gla.ac.uk/wiki/Main_Page
Organisms: Trypanosoma brucei
The aims of this investigation is to quantify metabolites associated with pathways involved in stress responses for parameterising models of oxidative stress metabolism; the measurement of metabolic fluxes of metabolites of interest with intracellular concentrations
Submitter: Dong-Hyun Kim
Studies: Metabolic flux measurement, Targeted metabolite analysis, Untargeted metabolite analysis
Assays: Generation of uniformly 13C-labelled E. coli extract, Intracellular metabolite concentrations in T. brucei exposed to oxidativ..., Intracellular metabolite concentrations in T. brucei under pH stress, LC-MS based absolute quantification of extracellular metabolites, LC-MS based absolute quantification of intracellular metabolites, Metabolite profiling on T. brucei exposed to oxidative stress
Snapshots: No snapshots
This assay is for method development to quantify intra- and extra-cellular metabolites on T. brucei 427 bloodstream form using isotope ratio based MS technique with 13C-labelled E. coli extract
Submitter: Dong-Hyun Kim
Assay type: Metabolite Concentration
Technology type: Mass Spectrometry
Investigation: Metabolite profiling, quantification and flux q...
Study: Metabolic flux measurement
Organisms: Trypanosoma brucei : Tbrucei427 (wild-type / wild-type)
SOPs: Generation of uniformly 13C-labelled E. coli ex...
Data files: No Data files
Snapshots: No snapshots
Intracellular metabolites in T. brucei at different stage of cell growth have been quantified absolutely by isotope ratio based MS technique using uniformly 13C-labelled E. coli extract. This is the case study for method development of absolute quantification for metabolic flux analysis.
Submitter: Dong-Hyun Kim
Assay type: Metabolomics
Technology type: Mass Spectrometry
Investigation: Metabolite profiling, quantification and flux q...
Study: Metabolic flux measurement
Organisms: Trypanosoma brucei : Tbrucei427 (wild-type / wild-type)
SOPs: Sample preparation for intracellular metabolite...
Data files: Absolute intracellular concentrations of metabo...
Snapshots: No snapshots
Extracellular metabolites in T. brucei at different stage of cell growth have been quantified absolutely by isotope ratio based MS technique using uniformly 13C-labelled E. coli extract. This is the case study for method development of absolute quantification for metabolic flux analysis.
Submitter: Dong-Hyun Kim
Assay type: Extracellular Metabolite Concentration
Technology type: Isotope Ratio Mass Spectrometry
Investigation: Metabolite profiling, quantification and flux q...
Study: Metabolic flux measurement
Organisms: Trypanosoma brucei : Tbrucei427 (wild-type / wild-type)
SOPs: Sample preparation procedure for footprint anal...
Data files: Concentrations of extracellular metabolites fro...
Snapshots: No snapshots
Intracellular concentrations of 50 metabolites measured by LC-MS using isotope ratio based mass spectrometry technique
Creator: Dong-Hyun Kim
Submitter: Dong-Hyun Kim
Investigations: Metabolite profiling, quantification and flux q...
Studies: Metabolic flux measurement
Concentrations of 22 extracellular metabolites (major medium components) from T. b. brucei 427
Creator: Dong-Hyun Kim
Submitter: Dong-Hyun Kim
Investigations: Metabolite profiling, quantification and flux q...
Studies: Metabolic flux measurement
Sample preparation procedure for metabolic analysis on T. b. brucei 427 using LC/MS
Creator: Dong-Hyun Kim
Submitter: Dong-Hyun Kim
Investigations: Metabolite profiling, quantification and flux q...
Studies: Metabolic flux measurement, Targeted metabolite analysis, Untargeted metabolite analysis
Assays: Intracellular metabolite concentrations in T. b..., Intracellular metabolite concentrations in T. b..., LC-MS based absolute quantification of intracel..., Metabolite profiling on T. brucei exposed to ox...
Labelling and extraction procedure for uniformly 13C-labelled E. coli (MG1655) for absolute quantification using isotope dilution technique by LC-MS
Creator: Dong-Hyun Kim
Submitter: Dong-Hyun Kim
Investigations: Metabolite profiling, quantification and flux q...
Studies: Metabolic flux measurement
Sample preparation procedure for metabolic footprint analysis on T. b. brucei 427 using LC/MS
Creator: Dong-Hyun Kim
Submitter: Dong-Hyun Kim
Investigations: Metabolite profiling, quantification and flux q...
Studies: Metabolic flux measurement