Cystoscopy is a diagnostic and invasive procedure for treatment and follow-up of genitourinary system patients and could be performed with a variety of anesthesia techniques. The study aimed to assess the efficacy of dexmedetomidine-ketamine vs. fentanyl-ketamine on sedation and analgesia for cystoscopy. This double-blind randomized controlled clinical trial enrolled 60 patients undergoing cystoscopy in two groups. Patients were assigned randomly by block random allocation method into dexmedetomidine-ketamine group (1 μg/kg dexmedetomidine) and fentanyl-ketamine group (2 μg/kg fentanyl) receiving ketamine (0.5 mg/kg). Subsequently, mean blood pressure, heart rate, saturated oxygen, respiratory rate, pain intensity, Ramsay score for sedation level, cystoscopy duration, and urologic satisfaction were measured and compared between two groups. Both the groups were similar regarding age, sex and baseline hemodynamic parameters (P > 0.05). Lower heart rate and pain score were revealed in the dexmedetomidine-ketamine group at 25-50 and 30-60 minutes, respectively, after cystoscopy (P < 0.05). Moreover, repeated measure test showed that there was significant difference in trend of respiratory rate and pain score between two groups (P = 0.017) and was lower in dexmedetomidine-ketamine group. The dexmedetomidine-ketamine group relieves pain 30 minutes after cystoscopy with stable hemodynamic parameters during operation. Therefore, dexmedetomidine-ketamine is recommended to be employed for pain relief in subjects undergoing cystoscopy. The study was approved by Ethical Committee of Arak University of Medical Sciences with IR.ARAKMU.REC.1397.108 on July 2, 2018, and registered in Iranian Registry Clinical Trial center with code IRCT20141209020258N105 on April 21, 2019.