Tunnel lay-by spacing is directly related to traffic safety and engineering investment. Nevertheless, its mechanism is not clear, and the rationality of the exiting norms with respect to tunnel lay-by spacing needs to demonstrate. A calculation model for tunnel lay-by spacing was derived by considering the headway and the physical kinematics of the two vehicles chasing and encountering. With it, the influence of various parameters on lay-by spacing were analysed and the rationality of the model were discussed through comparing with existing norms. Results shows longitudinal gradient rate, daily average traffic flow, rolling resistance coefficient, posted speed limit are significant to determine the lay-by spacing, and the most important parameter is longitudinal gradient rate. Existing tunnel lay-by spacing norm values are not reasonable enough, either too strict or too loose. These findings provide scientific support for how to select tunnel lay-by spacing value, which can improve tunnel traffic safety and make engineering investment reasonable.