The anesthetic effect of a combination of medetomidine, midazolam and butorphanol (Me-Mi-Bu) was evaluated in healthy cynomolgus monkeys. The Me-Mi-Bu combination was intramuscularly administered as follows: Dose 1, Me 0.015 mg/kg-Mi 0.1 mg/kg-Bu 0.15 mg/kg; Dose 2, Me 0.02 mg/kg-Mi 0.15 mg/kg-Bu 0.2 mg/kg; and Dose 3, Me 0.04 mg/kg-Mi 0.3 mg/kg-Bu 0.4 mg/kg. The combination rapidly induced immobilization, and lateral recumbency was reached within 15 min. The duration of anesthesia for each dose administered was follows: Dose 1, 47 ± 27 min; Dose 2, 113 ± 31 min; and Dose 3, 190 ± 24 min. The anesthetic effect of the combination was abolished by the α2-adrenoceptor antagonist atipamezole. No marked changes in the levels of hematologic or serum biochemical parameters were noted in cynomolgus monkeys administered the combination plus atipamezole. Taken together, these results suggest that the Me-Mi-Bu combination exhibits reversible anesthetic effect and may be useful for studies involving cynomolgus monkeys.