The main aim of the study was to investigate the differences in repeated sprint ability of first- compared to second-division referees at the beginning and in the middle of the soccer season. Twenty-seven main and 51 assistant referees, from the first (age 36.2 ± 5.3) and the second (age 28.8 ± 3.9) soccer divisions, performed two fitness tests—at the beginning and at mid-season, consisting of a repeated sprint test of 6 × 40 m and 5 × 30 m sprints for the referees and assistant referees, respectively. While no differences were found among the first division’s main referees, significant improvement ( p < .05) was found among the second division’s main referees at mid-season compared to the beginning of the season in ideal sprint time (31.97 ± 1.5 vs. 32.47 ± 1.3 s) and total sprint time (32.62 ± 1.4 vs. 33.10 ± 1.1 s), respectively. The main referees' ideal sprint time and total sprint time were significantly faster ( p < .05) in the second compared to the first division, at pre-season and at mid-season. Significant correlations were found between age and ideal sprint time ( r = .63) and age and total sprint time ( r = .66) among the main referees, but not between age and performance indices among the assistant referees. The findings demonstrated a superiority of the second over the first division's referees in anaerobic capabilities, suggesting that training program should be specific for each division. It may also indicate that variables other than physical capabilities determine referees' professional status and referee's advancement from second to the first division.