Abstract: Proximity-dependent biotinylation (PDB) uses biotin ligase fused to the protein of interest to biotinylate adjacent proteins, purify them with streptavidin beads, and then identify the biotinylated protein by mass spectrometry.This technology can be used to detect transient and/or low affinity interactions, provide a chance to learn more about membrane-less organelles and other subcellular structures that cannot be easily isolated or purified, and fill the gap in traditional methods.This article summarizes the technological development and application of PDB in recent years.