Neospora spp. and Toxoplasma gondii are two closely related protozoan parasites which are widely distributed through the world by causing fetal malformations and neurological disorders in a wide range of animals and human in T.gondii. Horses can act as intermediate hosts for both parasites and can acquire disease. Blood samples were taken from 487 clinically healthy horses of 17 different mechanized stables in Tehran, the capital of Iran, during September and November of 2022. Through sample collection, information regarding animal characteristics, local management and environmental conditions were recorded. IFAT and ELISA was employed to detect antibodies directed against Neospora spp. and T. gondii. The IFAT results show that anti-N. caninum antibodies were detected in 52 of 487 animals (10.67%) and antibodies against T. gondii were detected in 41 horses (8.42%). Also, In 6 of 487 horses (1.23%) double positivity suggested co-infection with both parasites. Gender, age groups, and the presence of dogs for neosporosis, and age groups and the presence of cats for toxoplasmosis, could be considered as factors having an influence on the seroprevalences (P<0.05). The results proved these parasites are present suggests that powerful regulatory measures should be implemented to prevent and control the spread.