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The TOP method is a simple, robust, environment friendly proteome extraction method, with decreased fixation time bias. Using the TOP method, we analyzed by LC\MS-MS a clinical cohort of microsatellite stable (MSS) and unstable (MSI-H) colorectal carcinoma (CRC). An MSI-H specific, STAT-1 centric, immunological signature was identified. In-vitro experiments connected this signature to long, but not short exposure, to Interferon-g. Our data provides in-depth view of the MSI-H immunobiology and suggests that the roles of STAT proteins are context dependent., Graphical Abstract Highlights • TOP: robust, bio-friendly FFPE proteome extraction method with less fixation bias. • Proteome of MSI-H colorectal cancer identifies immunobiology key elements. • MSI-H tumor displays an “INFg-STAT1 centric signature”. • Long-term IFNg induction In-vitro mimicks MSI-H signature., Using a simple, environment friendly proteome extraction (TOP), we were able to optimize the analysis of clinical samples. Using our TOP method we analyzed a clinical cohort of microsatellite stable (MSS) and unstable (MSI-H) colorectal carcinoma (CRC). We identified a tumor cell specific, STAT1-centered, immune signature expressed by the MSI-H tumor cells. We then showed that long, but not short, exposure to Interferon-γ induces a similar signature in vitro. We identified 10 different temporal protein expression patterns, classifying the Interferon-γ protein temporal regulation in CRC. Our data sheds light on the changes that tumor cells undergo under long-term immunological pressure in vivo, the importance of STAT proteins in specific biological scenarios. The data generated could help find novel clinical biomarkers and therapeutic approaches.