ICPL labeling strategies for proteome research.

Stable isotope labeling in combination with mass spectrometry has emerged as a powerful tool to identify and quantify thousands of proteins within complex protein mixtures. Isotope-coded protein label (ICPL) is capable of high-throughput quantitative proteome profiling on a global scale. Since ICPL is based on stable isotope tagging at the free amino groups of intact proteins, it is applicable to any protein sample, including extracts from tissues or body fluids. After labeling of up to four different proteome states, the samples can be combined and the complexity reduced by any separation method currently employed in protein chemistry. After enzymatic cleavage of the protein fractions the ratios of peptides in the different proteome states can be calculated by simple MS-based mass spectrometric analyses. Only peptides that exhibit regulations in the different proteome states are further investigated for identification by tandem-mass spectrometry. The quantification of multiplexed ICPL experiments is greatly facilitated by the recently published ICPLQuant software, which was especially designed to cover the whole ICPL workflow. The method shows highly accurate and reproducible quantification of proteins, yields high sequence coverage, and is indispensable for the comprehensive detection of posttranslational modifications and protein isoforms.