The biomechanics of steady-state locomotion in different breeds of working dog is understudied, despite widespread use of these animals in multiple industries. It is unknown how kinematic and kinetic parameters vary between breeds and how these variations are potentially related to inter-breed variations in morphology. Here, gross morphology and trotting locomotion within a cohort of 27 Labrador Retrievers (“Labradors”), shepherd breeds (“Shepherds”) and spaniel breeds (“Spaniels”) were compared using motion capture, force plates and biomechanical modelling. Evidence for slight positive allometric scaling of limb and body lengths was found between the breeds, with relatively longer lengths seen in Shepherds compared to Spaniels. Significant between-breed differences in raw spatiotemporal parameters were found with the larger Shepherds trotting with greater velocities, stride lengths and ground reaction forces than the smaller breeds, although many of these factors scaled with isometry with respect to body mass when accounting for variations in trotting speed. However, gait cycle times and stride lengths do not scale isometrically with body size, which, taken together with significant differences in flexion-extension joint angles and moments, suggest dynamic similarity during trotting is unlikely between these breeds. Overall, these findings highlight specific differences in the biomechanics of steady-state trotting locomotion between working dog breeds despite their somewhat geometrically similar gross body proportions. This suggests that not only should locomotion databases for individual breeds exist for future canine research, but also that breed-specific considerations should be adopted to maximise the health and welfare of these dogs in working practices, such as load-carrying tasks.