The reproductive systems of male and female Drosophila have been well studied genetically and cytologically because of the importance of meiotic mechanisms in understanding inheritance and speciation. Recently, there has been increasing interest in proteomics of Drosophila melanogaster, because detailed and comprehensive gene annotations have made it straightforward to identify proteins analyzed by mass spectrometry. We have applied proteomic analysis to the male reproductive system with the aim of understanding sperm maturation not only during the process of spermatogenesis, but also during the movement of sperm from the seminal vesicle through the male reproductive system and into the uterus following copulation. In this article, we will provide an overview of the principles of recent proteomic technologies and describe a few proteomic profiles of the male reproductive organs.