An in vivo labeling strategy with leucine-d3 for quantitative proteomics applied to the study of muscle cell differentiation

The mass spectrometric methods were used for the identification and quantitation of complex protein mixtures. The stable isotope labeling by amino acids (SILAC) method was used for the labeling by amino acids in cell culture for the in vivo incorporation of specific amino acids into all mammalian proteins. The mammalian cell lines were grown in media lacking a standard essential amino acid and the cultured cells specifically incorporated the stable isotope containing amino acids. The results show that the advantages of using stable isotopes to label proteins are, SILAC requires no peptide labeling steps and since the extent of incorporation is virtually 100% there are no differences in labeling efficiency between one sample and the other.