Interactome mapping, the systematic identification of protein interactions within an organism, promises to facilitate systems-level studies of biological processes. Using in vitro technologies that measure specific protein interactions, static maps are being generated that include many of the protein networks that occur in vivo. Most of the binary protein interaction data currently available was generated by large-scale yeast two-hybrid screens. Recent efforts to map interactions in model organisms and in humans illustrate the promise and some of the limitations of the two-hybrid approach. Although these maps are incomplete and include false positives, they are proving useful as a framework around which to elaborate and model the in vivo interactome.