AbstractCdc25C is a dual specificity phosphatase essential for dephosphorylation and activation of cyclin‐dependent kinase 1 (cdk1), a prerequisite step for mitosis in all eucaryotes. Cdc25C activation requires phosphorylation on at least six sites including serine 214 (S214) which is essential for metaphase/anaphase transit. Here, we have investigated S214 phosphorylation during human meiosis with the objectives of determining if this mitotic phosphatase cdc25C participates in final meiotic divisions in human oocytes. One hundred forty‐eight human oocytes from controlled ovarian stimulation protocols were stained for immunofluorescence: 33 germinal vesicle (GV), 37 metaphase stage I (MI), and 78 unfertilized metaphase stage II (MII). Results were stage dependent, identical, independent of infertility type, or stimulation protocol. During GV stages, phospho‐cdc25C is localized at the oocyte periphery. During early meiosis I (MI), phosphorylated cdc25C is no longer detected until onset of meiosis I. Here, phospho‐cdc25C localizes on interstitial microtubules and at the cell periphery corresponding to the point of polar body expulsion. As the first polar body reaches the periphery, phosphorylated cdc25C is localized at the junction corresponding to the mid body position. On polar body expulsion, the interior signal for phospho‐cdc25C is lost, but remains clearly visible in the extruded polar body. In atresic or damaged oocytes, the polar body no longer stains for phospho‐cdc25C. Human cdc25C is both present and phosphorylated during meiosis I and localizes in a fashion similar to that seen during human mitotic divisions implying that the involvement of cdc25C is conserved and functional in meiotic cells. Mol. Reprod. Dev. 75: 1176–1184, 2008. © 2007 Wiley‐Liss, Inc.