What is Mass Spectrometry?

Mass Spectrometry or MS is one of the advanced techniques that is used to systematically study and examine a large number of proteins in a cell, tissue or an organism. Recently, with advancement in the area of cutting edge proteomic technology, MS has become the primary tool for differential proteomic analyses, identification of protein biomarkers and understanding of extensive protein-protein interactions in cancer. By determining the molecular mass of charged peptide particles, mass spectrometer simultaneously identify and accurately quantify thousands of unique protein signatures in cancer samples; demonstrating to be a very powerful yet sensitive and accurate technique for proteomic analyses.
In Mass Spectrometry, protein components of a sample is detected and analysed according to their mass-to-charge (m/z) ratio in MS and/or MS/MS mode. In MS mode, the m/z ratio of precursor tryptic peptide units is analysed while in the MS/MS mode the m/z ratio analyses is extended to the ion fragments of the tryptic peptide units.
For MS-based proteomic analyses, the protein samples are first prepared and purified followed by their digestion with trypsin; an enzyme that cuts large and complex proteins into smaller peptides.
Often, Liquid Chromatography (LC) is coupled with mass spectrometers to separate the peptide mixtures in a liquid mobile phase on the basis of their interactions with the column’s stationary phase. Depending on their polarity and their size, different analytes will interact differently with both stationary and mobile phases. As the mobile phase moves along the column, the analytes will separate over time and elute from the column into the mass spectrometer at a time known as retention time. In this method, the highly complex peptide mixture is injected onto the LC column where they are separated in one or higher LC gradient. As peptides elute from the LC column, they enter the Mass Spectrometer where they are ionised followed by their detection and identification in MS and MS/MS mode.
Finally, the mass spectrometric raw data which contains both MS and MS/MS information will be analysed and searched against database for the identification and quantification of protein IDs using search algorithms such as Mascot Daemon, Proteome Discoverer and MaxQuant. Statistical analyses are then performed on the search result to evaluate the relative expression of proteins of interest in different tumour types, and their significance level are calculated.
By Ali Azimi