Techniques

Biosynthetic pathway analysis

Collaboration with the lipidomics research facility will yield powerful biological interpretation of the lipid profile contained in experimental mammalian samples. Pathway analysis bridge the gap between structurally-related lipids regulated by enzymes from well known metabolic pathways. Visual quantitative results (shown below) is provided with a list of genes, which could be involved in the activation or suppression of enzymes catalysing lipid metabolism in mammals. The quantity and detail of the quantitative lipidome results obtained by the facility is subjected to in-depth careful statistical analysis using R-studio (developed in collaboration with the bioinformatics group on-site) and MATLAB scripts.

 

Chromatography and mass spectrometry

Samples are subjected to a variety of liquid/liquid extractions, which are chosen depending on the lipid subclass to analyse (as shown in the figure below). Interestingly, processing/pelleting of samples in the early stages of sample collection can affect lipid recovery. Contact us to discuss your sample requirements to avoid differential loss of lipids by Email: lipidomics@babraham.ac.uk.

Lipid extracts are then injected into diverse chromatographic columns to offer orthogonality to mass spectrometry analysis. Chromatography is also a key step to separate abundant lipid subclasses/molecular species from low abundant moieties preventing ion supression of the low abundant signalling lipids.

The lipidomics research facility staff have actively developed eight chromatographic methods and includes the analysis of phosphoinositol mono, di, and triphosphate developed in the Biological Chemistry facility (doi:10.1038/nmeth.1564).

Lipidomics techniques pathway

Lipid short notation and nomenclature

Each lipid subclass molecular species identified using chromatography coupled with mass spectrometry is reported using the LIPID MAPS classification system and short hand notation. 40 lipid subclasses molecular species are identified and semi-quantified of a "before" sample and an "after" sample, and then comparisons of the data sets are made to see what has changed. All of these analysis form part of the collaborations established with the lipidomics research facility.

LipidMaps Classification System