Bottom-Up Mass Spectrometry–Based Proteomics as an Investigative Analytical Tool for Discovery and Quantification of Proteins in Biological Samples

Abstract

Objective: The objective of this overview is to introduce bottom-up mass spectrometry (MS)-based proteomics approaches and strategies, widely used in other biomedical research fields, to the wound-healing research community. Approaches: Two major proteomics workflows are discussed: gel-based and gel-free chromatographic separation to reduce the complexity of the sample at protein and peptide level, respectively, prior to nano-liquid chromatographytandem mass spectrometry analysis. Other strategies to discover less abundant proteins present in the sample, are also briefly discussed along with label-free and label-incorporated methods for protein quantification. Overall, the experimental workflows are designed and continually improved to increase the number of proteins identifiable and quantifiable. Discussion: Recent advances and improvements in all areas of proteomics workflow from sample preparation, to acquisition of massive amounts of data, to bioinformatics analysis have made this technology an indispensable tool for in-depth large-scale characterization of complex proteomes. This technology has been successfully applied in studies focusing on biomarker discovery, differential protein expression, protein-protein interactions, and post-translational modifications in complex biological samples such as cerebrospinal fluid, serum and plasma, and urine from patients. The publications from these studies have reported greater number of identified proteins, novel biomarker candidates, and post-translational modifications previously unknown. Conclusions: The qualitative and quantitative protein analysis of the protein population of wound tissues or fluids at different stages is important in wound healing research. Given the complexities and analytical challenges of these samples, MS-based proteomic workflows further improved with recent advances offer a powerful and attractive technology for this purpose.

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