Proteomics Analysis of the Estrogen Effects in the Rat Uterus Using Gel-LC and Tandem Mass Spectrometry Approaches.

Proteomics-based bottoms-up, at a big scale applied to the protein identification and relative quantification present in complex mixtures (cell lysates, tissues, biological fluids, secretome, etc.) is a useful strategy to identify proteins and analyze their changes. Samples processed through a gel-free approach provide a simple method for protein separation and profile comparison of different conditions, such as using fewer steps in the protocol, reducing excessive sample handling, and covering an extended range of molecular weights and isoelectric points. However, it presents a great limitation related to the management of large dynamic ranges of proteins. There are numerous protocols that allow handling the problem or limitations generated by a high dynamic range of the proteins present in the sample. The Gel-LC technique is a complementary alternative of the gel-free approach available to solve the issue of protein samples with a high dynamic range. The different steps of the protocol involve sample processing through Gel-LC (1D-SDS-PAGE) prior to digestion, 1D-nanoUHPLC coupled to high-resolution/mass accuracy tandem mass spectrometry analysis (1D-nanoUHPLC-HR/MA-MS /MS analysis) and afterward, the protein identification and relative quantification analysis using bioinformatics tools for the data conversion, organization, and interpretation.