The study aims were to describe positional differences in the acceleration and sprint profiles of professional football players in match-play, and analyse start speeds required based on the intensity of accelerations and decelerations. This longitudinal study was conducted over thirteen competitive microcycles in a professional football team from LaLiga 123. Data were collected through electronic performance tracking systems. Every player was categorised based on the playing position: central defender (CD), full-back (FB), forward (FW), midfielder (MF), and wide midfielder (WMF). In respect of acceleration profile, positional differences were found for all variables (p < 0.05), except average magnitude of accelerations (ACCAVG, p = 0.56) and decelerations (DECAVG, p = 0.76). The sprint profile also showed positional differences for all variables (p < 0.05), apart from sprint duration (p = 0.07). In addition, although low-intensity accelerations required significantly greater start speeds (Vo) than high-intensity accelerations in WMF (0.4 ± 0.2 km/h; p < 0.05) and FW (0.4 ± 0.2 km/h; p < 0.05), no significant differences (p > 0.05) were found in CD, FB, and MF. However, high-intensity decelerations were performed at significantly higher Vo than low-intensity decelerations in MF (2.65 ± 0.1 km/h; p < 0.05), FW (3.3 ± 0.1 km/h; p < 0.05), FB (3.9 ± 0.4 km/h; p < 0.05), WMF (4.3 ± 0.3 km/h; p < 0.05), and CD (4.1 ± 0.7 km/h; p < 0.05). Therefore, positional differences exist for most variables of the acceleration and sprint profiles. In addition, different Vo were observed between high-intensity and low-intensity accelerations as well as high-intensity and low-intensity decelerations.