Stable isotopic labelling is the most reliable and accurate method for quantitative proteomics and metabolomics. Proteomics studies on rats labelled uniformly with 15N have been previously reported (McClatchy et al. 2007) using 15N-labelled algae as protein source.
Silantes has developed in cooperation with the group of Prof. Chris Turck, Max-Planck-Institute of Psychiatry (Zhang et al. 2008), a 15N-diet specially designed for labelling mice uniformly using the hydrolysate of the chemolithoautotrophic bacteria, Ralstonia eutropha, as 15N-protein source. Feeding experiments showed that Ralstonia-based protein is better accepted by mice than the algae-based protein.
The concept in short: In order to determine the protein pattern of mouse (A) with respect to mouse (B), specimens of mice (A) and (B), both of which were fed a diet containing only unlabelled (14N)-protein (from Ralstonia), are each mixed with specimen of a reference mouse (R) which was fed a diet in which the 14N-protein has been replaced by 15N-protein (from Ralstonia).
Figure1 on the next page shows the workflow scheme of U-15N-SILAM, Figure 2 the feeding scheme and figure 3 the time dependent incorporation.
Figure 3 indicates that a 92% incorporation is achieved after about 56 days.
Silantes has developed a kit for performing the U-15N-SILAM-Mouse.
This U-15N-SILAM-Mouse Diet Kit consists of two feeds:
◾ U-14N-SILAM-Mouse Diet:
A feed (from Harlan) which is composed of a conventional protein-free diet. This diet is supplemented by a protein based on unlabelled (14N) Ralstonia hydrolysate (light feed). This diet is used for feeding mice (A) and (B);
◾ U-15N-SILAM-Mouse Diet:
The same feed as above but 14N-Ralstonia-based hydrolysate is replaced by 15N-Ralstonia-based hydrolysate (heavy feed). This diet is used for feeding the reference mouse (R).
Daniel B. McClatchy, Meng-Qiu Dong, Christine C. Wu, John D. Venable, and John R. Yates (2007). 15N metabolic labeling of mammalian tissue with slow protein turnover, Journal of Proteome Research, 6(5), 2005-2010.
YY. Zhang, M.D. Filiou, G. Chen, H. Heumann, C.W. Turck (2008). Biomarker discovery in a mouse model of trait anxiety using 15N Metabolic Labeling, Poster HUPO Amsterdam.