The KIT is a putative marker for differentiating spermatogonia in stallions

• The KIT is expressed in differentiating spermatogonia in stallions.
• The KIT expression was observed at all reproductive stages of stallions.
• The KIT is not expressed in Leydig or Sertoli cells.

Putative markers have been discovered and are used to identify and separate certain lineage of spermatogonia. The KIT is a marker for differentiating spermatogonial stem cells in several species including mice and goats. The objectives of this study were (1) to investigate reproductive stage-dependent KIT expression patterns in stallions and (2) to identify spermatogonia subpopulations expressing KIT in stallion testes. To achieve these objectives, testicular samples were obtained during routine field castration of stallions. The reproductive stage of the stallions was classified as pre-pubertal (<1 year, n = 3), pubertal (1–1.5 year, n = 4), post-pubertal (2–3 year, n = 6), or adult (4–8 year, n = 6). For immunohistochemistry, KIT was used at a dilution of 1:200. In the pre-pubertal and pubertal stage, most germ cells were immunolabeled with KIT. In the post-pubertal and adult stages, immunolabeling of KIT was evident in the germ cells attached to the basement membrane of the seminiferous tubules with exception of some spermatogonia. Co-immunolabeling with KIT and deleted in azoospermia like (DAZL) showed different co-staining patterns, including KIT only, both KIT and DAZL), or DAZL positive germ cell populations alone. The KIT was not immunolabeled in Sertoli or Leydig cells at any reproductive stages. The result of Western blot analysis verified the cross-activity of the KIT antibody with horse testes tissue. In conclusion, KIT appears to be expressed in differentiating spermatogonia, and may be used to identify and isolate differentiating germ cells from stallions.