BackgroundAlthough normal haematological and serum biochemical values for both pet and wild birds have been published, little information is available on the haematological and serum biochemical values in long‐legged buzzards (Buteo rufinus).ObjectivesThis is the first study that aimed to define reference values of haematological, biochemical parameters, and protein electrophoretic fractions of long‐legged buzzards in Iran.MethodsBlood samples were collected from 30 clinically healthy adult long‐legged buzzards of both sexes. Hematological, biochemical parameters, and protein electrophoretic fractions were measured. The mean and standard deviations were calculated.ResultsMean values for red blood cells, packed cell volume, haemoglobin, and white blood cells were 2.72 ± 0.60 ×106/μl, 39.10 ± 3.70%, 13.45 ± 1.30 g/dl, and 3.92 ± 1.39 ×103/μl, respectively. Mean values for biochemistry parameters were total protein 4.46 ± 1.27 g/dl, albumin 1.78 ± 0.55 g/dl, creatinine 0.54 ± 0.22 mg/dl, uric acid 7.81 ± 2.89 mg/dl, calcium 9.63 ± 2.22 mg/dl, phosphorus 4.31 ± 1.00 mg/dl, glucose 398.87 ± 96.90 mg/dl, blood urea nitrogen 10.46 ± 3.85 mg/dl, alkaline phosphatase 127.01 ± 1.46 IU/L, aspartate aminotransferase 262.22 ± 116.30 IU/L, and alanine aminotransferase 56.63 ± 27.85 IU/L. Mean values for serum protein fractions included pre‐albumin, albumin, α‐1 globulin, α‐2 globulin, β‐ globulin, and ϒ‐globulin was 0.20 ± 0.09, 2.35 ± 0.67, 0.28 ± 0.13, 0.32 ± 0.07, 0.62 ± 0.24, and 0.68 ± 0.53 g/dl, respectively.ConclusionThe reference data presented in this study can be used as health assessment values for veterinary laboratories and clinicians when developing release criteria for rehabilitated long‐legged buzzards.