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. 2021 Mar 26;11:608201. doi: 10.3389/fonc.2021.608201

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Copyright © 2021 Orfanou, Argyros, Papapetropoulos, Tseleni-Balafouta, Vougas and Tamvakopoulos

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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Experimental workflow for identification of candidate Breast Cancer (BC) biomarkers or drug targets using a strategy that combines a membrane enrichment protocol with GeLC-MS/MS technique. (A) Membrane enriched, cytosolic and whole cell lysate fractions from the breast cancerous MDA-MB-231 (MDA) and HCC-1954 (HCC) cell lines were isolated and analyzed by Western blot, using well established BC membrane targets (HER2 and EGFR), to verify the success of the membrane enrichment. (B) Membrane enriched fractions were then subjected to 1D SDS-PAGE gel electrophoresis, in-gel trypsin digestion, and tandem mass spectrometric analysis on a high resolution Orbitrap. Protein identification and quantification was performed using bioinformatic tools (Sequest database search). Cell, whole cell lysate fraction; cyt, cytosolic fraction; mem, membrane fraction.