A Brief Progress in Methods for Deciphering Protein–Protein Interaction Networks

Protein–protein interactions (PPIs) are fundamental regulators of cellular function and disease. Systematic mapping of the interactome is essential for identifying therapeutic targets and advancing drug design, a pursuit that has driven significant innovation to capture the spatiotemporal regulation of PPIs in vivo. This review summarizes this methodological revolution. We outline foundational, first-generation techniques—yeast two-hybrid and co-immunoprecipitation—which established frameworks for binary interaction mapping and static network generation, especially when integrated with mass spectrometry. The discussion then pivots to second-generation methods, including proximity-dependent labeling and advanced imaging, which enable the capture of PPIs within their native, dynamic cellular contexts. We provide a comparative analysis of these techniques, detailing their principles, strengths, and limitations. The review concludes with a practical framework for method selection and a perspective on emerging frontiers—such as spatial proteomics and single-cell interactomics—that are poised to further decode the evolving interactome. This concise overview serves as a strategic guide for specialists adopting new techniques and a broader audience integrating network-level data into their research.