Pseudo Internal Standard Approach for Label-Free Quantitative Proteomics

Quantitative proteome analysis has become a versatile tool to understand biological functions. Although stable isotope labeling is the most reliable method for quantitative mass spectrometry, preparation of isotope-labeled compounds is time-consuming and expensive. Simple label-free approaches have been introduced, but intensity-based quantitation without standards is not generally accepted as reliable, especially for small molecules. We have developed a novel label-free quantitative proteome analysis using pseudo internal standards (PISs). This idea was derived from northern blotting analysis, in which housekeeping genes are used as standards to normalize and compare target gene expression levels in different samples. In many proteomics studies, most proteins do not change their expression levels under different conditions, and therefore, these proteins can be employed as pseudo internal standards. This new approach is simple and does not require additional standards or labeling reagents. The PIS method represents a novel approach for mass spectrometry-based comprehensive quantitatitation and may also be applicable to quantitative metabolome analysis.