Quantitative Analysis of chemically modified biopolymers using separation techniques and mass spectrometry

Modern proteomic research calls not only for qualitative analysis of peptides and proteins but also for quantitative analysis. Only the quantity of proteins as well as its post-translational modifications (PTMs) or a change in these quantities could provide deeper insights into the status of biological systems, which could be related to the various normal and disease states. Furthermore, the quest for significant biological markers that indicate tumours and other diseases requires sensitive, robust and reliable quantitative data. However, quantitative proteomics encounters challenges associated with sample complexity, wide dynamic range of protein abundance and different physico-chemical properties of proteins, which complicate the handling and analysis of the sample. The technological advances, particularly in separation techniques and mass spectrometry (MS) detection, allowed the development of several approaches to comprehensively assess proteomes for analysis. The separation techniques are mainly one- and two-dimensional polyacrylamide gel electrophoresis (1D or 2D-PAGE), which are powerful techniques in this respect, but in quantitative aspects they suffer from a lack of sensitivity and reproducibility, sample loss and low dynamic range. Therefore, 2D-PAGE was largely replaced by liquid chromatography (LC) techniques and LC coupled to molecule specific MS has emerged as a feasible alternative for qualitative characterization of complex mixtures.