The aim of this study was to investigate the thickness of acellular extrinsic fibre cementum (AEFC) at four root positions of anterior and posterior teeth with special focus on functional aspects. Furthermore, the correlations between cementum thickness and chronological age and sex are investigated. While numerous studies confirm continuous cementum apposition with age, masticatory forces as well as physiological and orthodontically induced tooth movements also have the potential to affect tooth cementum thickness. Undecalcified teeth were embedded in resin and transverse-sectioned in the cervical third of the root. Two sections per root were selected, and digital images at four positions were obtained (mesial, distal, oral, and vestibular) using light microscopy. The AEFC thickness of 99 teeth (anterior = 66, posterior = 33, male = 54, female = 45) were measured in both sections. The differences in mean values between root positions and the association of root position variation with tooth type, age, sex, and subject as well as the overall effects of age and sex were analysed using a mixed model. First incisors and canines showed the greatest mean AFEC thickness, in contrast to premolars which had the lowest values. Differences were found across the four root positions, with a pattern varying considerably between anterior and posterior teeth and between maxilla and mandible in the anterior teeth. An interaction between root position and subject pointed to the existence of an individual component in the variation of AEFC thickness across the four root positions. There was an age trend with an almost linear increase in cementum thickness of 1 μm per year. Overall, females tended to exhibit a significantly lesser AEFC thickness compared to males. Distinct differences in the pattern of thickness values across the four root positions in anterior and posterior teeth support the assumption that the AEFC is strongly affected by functional processes. In addition to sex-specific differences and age-related trends, the root position variation of AEFC thickness varies from individual to individual.