Abstract: Nanoindentation is an attractive characterization technique, as it not only measures the local properties of a material but also facilitates understanding of deformation mechanisms at submicron scales. However, because of the complex stress–strain field and the small scale of the deformation under the nanoindenter, the results can be easily influenced by artifacts induced during sample preparation. In this work, a systematic study was conducted to better understand the influence of sample preparation methods on the nanoindentation results of ductile metals. All experiments were conducted on a steel (Fe–22Mn–0.65C, wt%) with twinning-induced plasticity (TWIP), which was selected for its large grain size and sensitivity to different surface preparation methods. By grouping the results obtained from each nanoindent, chemical polishing was found to be the best sample preparation method with respect to the resulting mechanical properties of the material. In contrast, the presence of a deformation layer left by mechanical polishing and surface damage induced by focused ion beam (FIB) scanning were confirmed by the dislocation-nucleation-induced pop-in events of nanoindentation.