The maximal lactate steady state (MLSS) has been considered the gold standard method to determine aerobic/anaerobic metabolism transition during continuous exercise executed by human beings and rats. Therefore, the aim of the present study was to determine MLSS for aerobic evaluation in swimming mice. Twenty-five adult male mice (90-day-old animals) were adapted to the deep aquatic environment at the temperature of 31 ± 1°C. The mice were submitted to five continuous exercise loads of 3, 4, 5, 6 and 7% of the body weight (bw) tied to the back, executed with 25-min duration and 48-h intervals between them. Blood samples were collected from the tail during swimming exercise (rest, 5, 10, 15, 20 and 25 min) for blood lactate concentration (LAC) determinations. The individual MLSS was considered as the highest intensity in which the increase on the LAC was equal to or below 1 mmol l− 1 from the 10th to the 25th minute of exercise. The results showed that 36% of the swimming mice presented MLSS at 4% bw, 20% at 3% bw and 6% bw, 16% at 5% bw and 8% at 7% bw. The LAC at the MLSS was 5.78 ± 0.29 mmol l− 1 (4.40–6.67 mmol l− 1). These results indicate that the MLSS of mice swimming with additional weight for the final 15 of 25 min of exercise could be determined.