Introduction: With the worldwide decline on male fertility potential, the importance of the insight of the spermatogenic process has been increasing. In recent years, proteomic methodologies have been applied to seminal fluid of infertile men to search for infertility potential biomarkers. However, to understand the spermatogenic event and to search for treatment to spermatogenic impairment, comparative analysis of testicular proteomics is considered a powerful methodology. Areas covered: Herein, we present a critical overview of the studies addressing proteomic alterations in the development of spermatogenesis during puberty, as well as during the different phases of the spermatogenic event. The comparative studies of the proteomic testicular profile of men with and without spermatogenic impairment are also discussed and key proteins and pathways involved highlighted. Expert opinion: The usage of whole human testicular tissue with its heterogeneous cellular composition makes proteome data interpretation particularly challenging. This may be minimized by controlled experiments involving the collection of testicular tissue and sperm from the same individuals, integrated in a clinically characterized cohort of healthy and infertile men. The analysis of specific subcellular proteomes can add more information to the proteomic puzzle, opening new treatment possibilities for infertile/subfertile men.