Introduction: Factors affecting hatchability are mentioned as important indicators in breeder flocks’ performance, one of these elements is fertility. Roosters’ as half part of fertility have great role and aging reduces their fertility. The biologic system in live organisms such as animals is under permanent attack due to natural consequence of the body’s normal metabolic activity that produces free radicals. It has been revealed that phospholipid fraction of avian spermatozoa membranes has high proportion of polyunsaturated fatty acids (PUFA) and it is the reason why the spermatozoa are susceptible to free radical attack. During evolution, to deal with Reactive Oxygen Species (ROS), living organisms have developed specific antioxidant protective mechanisms. Therefore, as a major factor, presence of natural antioxidants in living organisms enables their survival in an oxygen-rich environment. To maintain sperm fertilizing ability, an antioxidant defense system is a crucial point. In avian semen the antioxidant system consists of natural antioxidants together with enzymes that have antioxidant characteristics such as glutathione peroxidase (GPx) and selenoprotein P (SEPP1) protects sperm against free radicals and their destructive metabolites. Antioxidants such as vitamin E and selenium (Se) have remarkable roles in avian reproduction. To gain gross reproductive performances in commercial poultry, supplementation of antioxidant at an optimum level is thought to be necessary. By using additives such as selenium we can help delaying this reduction through antioxidant properties of Se. Replacing inorganic Se by new types of Se sources like the nano form in poultry diet improves fertility. This research was conducted to investigate the effect of Nano-Selenium (Nano-Se) in comparison with sodium selenite on serum selenoprotein p (SEPP1) and glutathione peroxidase (GPx) content in broiler breeder roosters.Material and Method: In the present study, a total of thirty Arbor Acres Broiler Breeder roosters (40 weeks old) were randomly divided into five experimental groups. Each of which included 3 replicates of 2 birds. Relevant catalog recommended distribution of 160 gr of diet per rooster per day for broiler breeder males. After one-week adaptation they were fed with the basal diet (T1) supplemented with 0.3 mg/kg sodium Selenite (T2), 0.15 mg/kg nano-Se (T3), 0.3 mg/kg nano-Se (T4) and 0.6 mg/kg nano-Se (T5). The duration of feeding experiment was four weeks. At the end of adaptation week, blood samples were collected (2.5 ml) and after separating the serum, they were stored in -20°C refrigerator. Another sampling was two weeks later in the middle of experiment that again blood samples were collected and their serum were stored at -20°C. Four weeks after the treatment was done at the end of experiment, the roosters were humanely euthanized by cervical dislocation, the 3rd and last sampling was implemented at the end of experiment that as in the previous sampling was done, the blood samples were centrifuged and separated serum was stored in -20°C. Then serum concentration of the antioxidant “SEPP1” was measured by ELISA method and “GPx” was analyzed using a spectrophotometry kit.Results and discussion: The values of SEPP1 and GPx serum concentration obtained at the first time sampling showed no significant differences between all groups (P < 0.05). Serum SEPP1 concentration examination in the second sampling showed that roosters fed with both mineral and nano form of selenium supplements, had significantly higher concentrations of selenoprotein P in their serum, compared to the control group (P < 0.05) which the highest concentration was related to the treatment of 0.6 mg/kg nano-Se (T5) in their diet. Among the two groups of treatment 2 and treatment 3, which were fed 0.3 mg/kg sodium selenite (T2) and 0.15 mg/kg nano-Se (T3), respectively, treatment 3 had higher concentration. The results of examining the SEPP1 serum concentration for the final time at the end of the study indicated that there is a process similar to the trend of changes in the serum concentration of selenoprotein P that was seen in the second sampling step. The only difference observed in serum concentration results of the second sampling compared to third time was that the difference between the T2 and T3 treatments was greater in the last sampling. Simultaneously with SEPP1 analysis, serum glutathione peroxidase concentration was measured in three sampling times. At the second time sampling of serum GPx concentration, evaluation of the serum concentration of glutathione peroxidase revealed that the serum concentrations of this selenoprotein in broiler breeder roosters in groups T5, T4, T3 and T2 were significantly (P < 0.05) higher relative to its concentration in control group (T1). The difference between serum concentration of control group and T3 treatment was not significant (P < 0.05), also the difference in serum concentration of this selenoprotein between treatments T4 and T5 was not significant (P < 0.05). At the end of the study, after the third sampling and evaluation, the results of GPx serum concentrations revealed that all treatments showed a significant increase compared to the control group (P < 0.05) and with increasing the amount of nano-Se in the diet from 0.15 mg/kg to 0.6 mg/kg nano-Se, serum concentrations of glutathione peroxidase showed an increasing trend. Based on a consideration of all experiment indexes, in this research 0.6 mg/kg is suggested to be the best level of supplementation of nano-Se, and nano-Se showed higher contents of serum SEPP1 and GPx at the same amounts of nano-Se and sodium selenite supplementation. In conclusion, dietary supplementation of nano-Se was more effective than sodium selenite on serum SEPP1 and GPx concentration of tested selenoproteins in broiler breeder males.