Tektins (TEKTs) are constitutive filamentous proteins of microtubules in cilia, flagella, basal bodies, and centrioles. In mammals, five TEKTs (TEKT1, 2, 3, 4, and 5) have been identified in testis and spermatozoa. With the exception of TEKT1, these TEKTs have been reported to be present in spermatozoa with predominant localization at the peri-axoneme structures of flagella, i.e., mitochondria and outer dense fibers. In the present study, we produced an antibody against TEKT1 to examine the localization of TEKT1 in mouse, bull, and rat spermatozoa. By immunoblot analyses and immunofluorescence microscopy, we found TEKT1 to be present in sperm flagella and at the apical region of acrosome cap in spermatozoa of all these species. Acrosome-associated TEKT1 disappeared after in vitro acrosome reaction in mouse spermatozoa. These observations suggest another potential role for TEKT1 as a cytoskeletal element in the sperm head, or as a molecule involved in acrosome-related phenomena, such as acrosome reaction.