This study aimed to assess the variations in serum Anti-müllerian hormone (AMH) levels during the estrus and diestrus phases of the estrous cycle and its effect on fertility in cross-bred Hamdani ewes. In the study, 21 non-prolific cross-bred Hamdani ewes (aged 2-3 years with BCS 2.75-3.25) were subjected to estrous synchronization using intra-vaginal sponges. Following synchronization, ewes underwent natural mating and pregnancy examination was carried out on the 45^th day after mating through trans-rectal ultrasonography. Jugular blood samples were collected on day of estrus and 8 days later and analyzed for serum AMH and progesterone concentrations. Results showed that serum AMH levels were 100-480 and 95-520 pg/ml during estrus and diestrus phases, respectively. There was no change in serum AMH levels due to age, BCS and during estrus and diestrus phases of the estrous cycle (P>0.05). Estrus and diestrus serum AMH levels were significantly higher in pregnant ewes compared to non-pregnant ewes (P<0.05). Receiver-operating characteristic (ROC) analysis revealed that serum AMH cut-off value of >270 pg/mL was feasible to predict fertility in cross-bred Hamdani ewes. In conclusion, serum AMH can be used as a marker of fertility in non-prolific cross-bred Hamdani ewes.