Diagnosis of tenosynovitis is usually confirmed on the basis of synovial fluid analysis (such as cytological and biochemical examinations). This study was designed to examine biochemical (serum amyloid A (SAA), haptoglobin (Hp) and D-dimer) and cytological (total nucleated cell count (TNCC), neutrophil, lymphocyte, and monocyte count) characteristics of serum and synovial fluid of the equine digital flexor tendon sheath (DFTS). Synovial fluid samples were aseptically collected in EDTA from 43 limbs in 20 horses with (study group) and 8 limbs in 8 horses without (control group) distention of DFTS and serum samples were collected from each horse. Lymphocyte, monocyte, neutrophil, and TNCC in the synovial fluid were statistically higher in the distended sheath that shows an inflammatory nature of the distention, however, concentrations of D-dimer was lower in serum (0.1, 0.10-3.80) and (0.2, 0.10-0.20) than synovial fluid (19.2, 17.78-20.00) and (20, 19.90-20.00)) in both groups. The serum SAA concentrations of the control group (1.7, 0.10-2.16) were significantly higher than the study group (1, 0.86-1.05) and the concentration of SAA in serum was higher than synovial fluid (0.89, 0.86-0.98) in the control group. Results of this current study show that a cytological evaluation of the synovial fluid is more valuable than biochemical findings in the diagnosis of the inflammatory nature of this condition.