Relative quantification methods isotopic approach

As the name suggests, label-free quantification does not require the incorporation of an isotopic label into peptides for quantitation of relative concentrations. Each sample is separately prepared, subjected to individual LC/MS runs, and quantification is performed through analysis of their mass spectra. There are two approaches used for label-free quantification. The first approach is based on the assumption that peptide/protein levels follow a linear relationship with area under the curve of the ion spectra. This method involves quantitation of relative levels by comparing the peak intensity of ions from multiple peptides. But experimental variations in sample preparation and... 

One of the main problems to overcome in order to realise quantitative proteomics is that mass spectrometry is not strictly a quantitative technique with respect to molecular abundance. One approach to overcome this problem is to perform comparative experiments in parallel where one sample is labelled with an isotope (such as 0 or H) and the other is unlabelled. Mass differences between labelled and unlabelled samples may be observed in a mass spectrometer and the relative ratios quantified (Figure 9.21). Obviously this method of quantification relies on equivalent mass expression by both samples in spite of the isotopic labelling of the one and not the other. In other words, in spite of the labelling, both samples should behave identically chemically. The simplest way to effect labelling is to use isotope-coded affinity tags (ICATs). According to this technique, two identical populations of proteins... 

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