After ejaculation, mammalian spermatozoa are not capable of fertilizing a metaphase II-arrested egg. They require to undergo a series of biochemical and physiological processes collectively known as capacitation. In all these processes, the regulation of calcium ions fluxes plays essential roles and involves participation of many channels and transporters localized in the plasma membrane as well as in the membrane of intracellular organelles. In mammalian sperm, a fraction of these molecules has been proposed to contribute to mature sperm function. However, in many cases, the evidence for the presence of a given protein is based on the use of agonists and antagonists with more than one target. In this review, we will critically analyze the published evidence supporting the presence of these molecules in mammalian sperm with special emphasis to methods involving tandem mass spectrometry identification, electrophysiological evidence and controlled immunoassays.