Sample preparation, quenching and metabolite extraction
- Quenching and liquid/liquid metabolite extraction from adherent cells
- Quenching and direct extraction of metabolites from adherent cells
- Quenching of bacteria in solution (NaCl/Ethanol)
- Quenching eukaryotic suspension cells
- Isolation of mitochondria from cell lines
GC-MS usage
- general rules
- Preparing the machine and adding your samples
- Qualitative analysis
- Quantitative analysis
- Derivatization with methoxyamine and MSTFA
- GC-MS analysis of polar metabolites
- "Saponification" and methylation of fatty acids derived from lipids
- GC-MS analysis of fatty acid methylesters from cellular extracts
- GC-MS analysis of fatty acid methyl esters a la Joseph
LC-MS usage
- general rules for Agilent 6550 system (french and english)
- Checklist before running your samples (french and english)
- Preparing the machine and adding your samples
- Quantitative analysis
- Qualitative analysis
- Screening for unexpected changes of known or unknown metabolites
HPLC
- HPLC for nucleotides on SAX column
- Determination of H2S and thiosulfate by HPLC
Determination of phosphate release
- Fiske-Subbarow
- Molybdate-Malachite Green method (Itaya)
- Mild conditions (Saheki)
Thin layer chromatography:
Determination of metabolites and enzymatic activities
Manipulation of metabolites
- Desalting metabolites by gel filtration
- Reduction of carbonyl groups with Sodium borohydride/-deuteride