Fig. 3
From: Tumor secretome shapes the immune landscape during cancer progression
Overview of the main methods for detecting tumor secretome in research. (A) ELISA. Fluids are collected and placed on ELISA plates that have been pre-coated with a specific capture antibody. The target protein adheres to this antibody, and then an enzyme-linked detection antibody is introduced. The protein’s concentration is determined by measuring the colorimetric shift that occurs when the enzyme reacts with its substrate. (B) Antibody arrays. Samples containing secreted proteins from cell culture or body fluids are incubated with precoated antibody arrays. Specific antibodies immobilized on the array capture various secreted proteins. Bound proteins are detected using a reporter molecule conjugated to a secondary antibody, allowing simultaneous quantification of multiple proteins in a single sample and providing a comprehensive secretome profile. (C) Mass spectrometry after immunoprecipitation. After collection and concentration of cell culture supernatants or body fluids, secretome immunoprecipitation is performed. Biomolecules of interest are isolated and analyzed by mass spectrometry to identify and quantify secreted proteins and metabolites. (D) ER-BioIDHA biotinylation technique. The ER-BioIDHA plasmid is transfected into tumor cells to enable biotinylation of secreted proteins. Biotin is added to the culture and secreted proteins are biotinylated in situ. Streptavidin-based immunoprecipitation is performed to isolate the biotinylated proteins, which are then analyzed by mass spectrometry to identify a specific set of secreted molecules