The benefits of exposure to seminal plasma have been well demonstrated in vitro, but the ability of this substance to regulate sperm function and fertility in vivo is less understood. Utilising cauda epididymal ram spermatozoa as a model, with or without exposure to seminal plasma, our group has utilised novel live in vivo cell imaging, quantitative proteomics, carbohydrate profiling, neutrophil binding and artificial insemination trials to study the changes that occur to spermatozoa as they undergo the biological event of ejaculation. Comparative proteomic analysis of epididymal and ejaculated spermatozoa has shown that despite the complexity of the seminal plasma proteome a strikingly small number of unique proteins (BSP1, BSP5, EDIL3 & LEG1) are added to the sperm membrane as a consequence of ejaculation. Nonetheless, exposure to seminal plasma causes significant change in the relative amounts of surface sugars and provides a robust mechanism to protect spermatozoa against phagocytosis by neutrophils. We postulate these changes to the sperm glycocalyx could alter the visibility of sperm cells to the female immune system and explain how seminal plasma facilitates sperm transit through the female tract, particularly the cervix. Research is now focused on identifying the role of individual seminal plasma proteins, such as Binder of Sperm (BSP), in sperm function and testing their ability to support cryosurvival and navigability of the female tract. In time, it is hoped that physiologically relevant seminal plasma proteins can be utilised to improve the success of assisted reproductive technologies in a variety of species.