Pathogens are known to release in their environment a large range of toxins and other virulence factors. Their pathogenicity relies on this arsenal of exoproteins and their orchestrated release upon changing environmental conditions. Exoproteomics aims at describing and quantifying the proteins found outside of the cells, thus takes advantage of the most recent methodologies of next-generation proteomics. This approach has been applied with great success to a variety of pathogens increasing the fundamental knowledge on pathogenicity. In this chapter, we describe how the exoproteome should be prepared and handled for high-throughput identification of exoproteins and their quantitation by label-free shotgun proteomics. We also mentioned some bioinformatics tools for extracting information such as toxin similarity search.