The navigation accuracy of a star sensor depends on the estimation accuracy of its optical parameters, and so, the parameters should be updated in real time to obtain the best performance. Current on-orbit calibration methods for star sensors mainly rely on the angular distance between stars, and few studies have been devoted to seeking new calibration references. In this paper, an on-orbit calibration method using singular values as the calibration reference is introduced and studied. Firstly, the camera model of the star sensor is presented. Then, on the basis of the invariance of the singular values under coordinate transformation, an on-orbit calibration method based on the singular-value decomposition (SVD) method is proposed. By means of observability analysis, an optimal model of the star combinations for calibration is explored. According to the physical interpretation of the singular-value decomposition of the star vector matrix, the singular-value selection for calibration is discussed. Finally, to demonstrate the performance of the SVD method, simulation calibrations are conducted by both the SVD method and the conventional angular distance-based method. The results show that the accuracy and convergence speed of both methods are similar; however, the computational cost of the SVD method is heavily reduced. Furthermore, a field experiment is conducted to verify the feasibility of the SVD method. Therefore, the SVD method performs well in the calibration of star sensors, and in particular, it is suitable for star sensors with limited computing resources.