Protein Extraction From FFPE Kidney Tissue Samples: A Review of the Literature and Characterization of Techniques.

Most tissue biopsies from patients in hospital environments are formalin-fixed and paraffin-embedded (FFPE) for long-term storage. This fixation process produces a modification in the proteins called "crosslinks", which improves protein stability necessary for their conservation. Currently, these samples are mainly used in clinical practice for performing immunohistochemical analysis, since these modifications do not suppose a drawback for this technique; however, crosslinks difficult the protein extraction process. Accordingly, these modifications make the development of a good protein extraction protocol necessary. Due to the specific characteristics of each tissue, the same extraction buffers or deparaffinization protocols are not equally effective in all cases. Therefore, it is necessary to obtain a specific protocol for each tissue. The present work aims to establish a deparaffinization and protein extraction protocol from FFPE kidney samples to obtain protein enough of high quality for the subsequent proteomic analysis. Different deparaffination, protocols and protein extraction buffers will be tested in FFPE kidney samples. The optimized conditions will be applied in the identification by LC-MS/MS analysis of proteins extracted from 5, 10, and 15 glomeruli obtained through the microdissection of FFPE renal samples.

Proteomics analysis of the peritoneal dialysate effluent reveals the presence of calcium-regulation proteins and acute inflammatory response.

BACKGROUND: Peritoneal dialysis (PD) is a form of renal replacement used for advanced chronic kidney disease. PD effluent holds a great potential for biomarker discovery for diagnosis and prognosis. In this study a novel approach to unravelling the proteome of PD effluent based-on dithiothreitol depletion followed by 2D-SDS-PAGE and protein identification using tandem mass spectrometry is proposed. RESULTS: A total of 49 spots were analysed revealing 25 proteins differentially expressed, among them many proteins involved in calcium regulation. CONCLUSIONS: Remarkably, a group of proteins dealing with calcium metabolism and calcium regulation has been found to be lost through peritoneal dialysate effluent, giving thus a potential explanation to the calcification of soft tissues in patients subjected to peritoneal dialysis and kidney injury. Comparison of literature dealing with PD is difficult due to differences in sample treatment and analytical methodologies.