The objective of this study was to evaluate the response of buffaloes to automatic milking, examining the relationships between milking interval, milk production, and milking time for this species. A total of 7,550 milking records from an average of 40 buffaloes milked by an automatic milking system (AMS) were analyzed during a 3-mo experimental period at a commercial farm with Italian Mediterranean buffaloes in southern Italy. Date and time of animal identification, milk yield, milking duration, milking interval, and average milk flow rate were determined for each milking. The results were also used to predict the maximum number of milkings per day and the optimal number of buffaloes per AMS for different levels of milk production. The average interval period between 2 consecutive milkings was 10.3 h [standard deviation (SD) 3.3]. Overall, 3.4 and 25.7% of the milkings had an interval of ≤6 h or >12 h, respectively. Milking duration averaged 8.3 min per buffalo per milking (SD 2.7). The average milk flow rate was 1.3kg/min (SD 0.5) at a milk yield of 2.8kg per milking (SD 1.4). Assuming that the milking station is occupied 80% of the time, the number of milkings ranged from 136 to 152 per day and the optimal number of buffaloes per AMS ranged from 59 to 66 when the production level increased from 2 to 5kg of milk per milking. Automatic milking systems are suitable for buffalo, opening new options for the management of dairy buffalo farms.