Protocol for Increasing the Sensitivity of MS-Based Protein Detection in Human Chorionic Villi
| Autor: | Timur Shkrigunov, Pavel Pogodin, Victor Zgoda, Olesya Larina, Yulia Kisrieva, Maria Klimenko, Oleg Latyshkevich, Peter Klimenko, Andrey Lisitsa, Natalia Petushkova |
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| Jazyk: | angličtina |
| Rok vydání: | 2022 |
| Předmět: | |
| Zdroj: | Current Issues in Molecular Biology, Vol 44, Iss 5, Pp 2069-2088 (2022) |
| Druh dokumentu: | article |
| ISSN: | 1467-3045 1467-3037 |
| DOI: | 10.3390/cimb44050140 |
| Popis: | An important step in the proteomic analysis of missing proteins is the use of a wide range of tissues, optimal extraction, and the processing of protein material in order to ensure the highest sensitivity in downstream protein detection. This work describes a purification protocol for identifying low-abundance proteins in human chorionic villi using the proposed “1DE-gel concentration” method. This involves the removal of SDS in a short electrophoresis run in a stacking gel without protein separation. Following the in-gel digestion of the obtained holistic single protein band, we used the peptide mixture for further LC–MS/MS analysis. Statistically significant results were derived from six datasets, containing three treatments, each from two tissue sources (elective or missed abortions). The 1DE-gel concentration increased the coverage of the chorionic villus proteome. Our approach allowed the identification of 15 low-abundance proteins, of which some had not been previously detected via the mass spectrometry of trophoblasts. In the post hoc data analysis, we found a dubious or uncertain protein (PSG7) encoded on human chromosome 19 according to neXtProt. A proteomic sample preparation workflow with the 1DE-gel concentration can be used as a prospective tool for uncovering the low-abundance part of the human proteome. |
| Databáze: | Directory of Open Access Journals |
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