Instantaneous undeformed chip thickness is one of the key parameters in modeling of micro-milling process. Most of the existing instantaneous undeformed chip thickness models in meso-scale cutting process are based on the trochoidal trajectory of the cutting edge, which neglect the influences of cutter installation errors, cutter-holder manufacturing errors, radial runout of the spindle and so forth on the instantaneous undeformed chip thickness. This article investigates the tooth trajectory in micro-milling process. A prediction model of radial runout of cutting edge is built, with consideration of the effects of the extended length of micro-milling cutter and the spindle speed. Considering the effects of cutting-edge trochoidal trajectory, radial runout of cutting edge and the minimum cutting thickness, a novel instantaneous undeformed chip thickness model is proposed, and the phenomenon of single-tooth cutting in micro-milling process is analyzed. Comparisons of cutting forces under different chip thickness models and experimental data indicate that this new model can be used to predict cutting forces.