Neurofilaments are transported along axons stochastically in a stop-and-go manner, cycling between brief bouts of rapid movement and pauses that can vary from seconds to hours in length. Presently the only way to analyze neurofilament pausing experimentally on both long and short time scales is the pulse-escape method. In this method, fluorescence photoactivation is used to mark a population of axonal neurofilaments and then the loss of fluorescence from the activated region due to neurofilament movement is monitored by time-lapse imaging. Here we develop a mathematical description of the pulse-escape kinetics in terms of the rate constants of a tested mathematical model and we show how this model can be used to characterize neurofilament transport kinetics from fluorescence photoactivation pulse-escape experiments. This combined experimental and computational approach is a powerful tool for the analysis of the moving and pausing behavior of neurofilaments in axons.