In an in-depth study of the spermatozoon, the understanding of human physiology and pathology of the sperm epididymal maturation cannot be ignored. In this chapter, the epididymal transit of spermatozoa and the role of seminal plasma markers in predicting sperm quality and fertilization outcome are discussed. In the last years, the interest in studying the role of seminal plasma in male infertility is emerging. It is well established that routinely assessed semen parameters offer valuable insights into an individual’s fertility status; however, they lack both diagnostic and prognostic power. In this context, the diagnostic potential of seminal plasma presents promising opportunities for the identification of novel biomarkers to enhance the diagnosis of male infertility and to better predict fertilization outcomes.