Despite the efforts for precise alignment of satellite-based imaging sensors before launch, several factors may cause the values of the calibration parameters to vary between the time of ground calibration and on-orbit operation. This paper considers the problem of satellite-based imaging sensors on-board calibration while estimating the position and velocity of a target of opportunity. The pixel measurements (estimated location of the target’s image in the focal-plane array) generated by these sensors are used to estimate the sensors’ pointing angle biases, which is a key element of accurate tracking of a target in a space-based system. The target is assumed to be seen by the sensors from a changing direction as a function of the target position, allowing the target in this nonlinear tracking system to be observable. The evaluation of the corresponding Cramér–Rao lower bound on the co-variance of the bias estimates and the statistical tests on the results of simulations show that both the target trajectory and the biases are observable and that this method is statistically efficient.