Nanodiscs are self-assembled 10 nm particles composed of lipid bilayer patches, stabilized by helical amphipathic belt proteins. The size, monodispersity and well-defined structure make the nanodiscs a popular model for the biological cell membrane, especially for structural and functional studies of membrane proteins. The structures and properties of nanodiscs made of zwitterionic lipids are well known. However, the biological cell membrane is negatively charged and thus nanodiscs containing anionic lipids should provide a better mimic of the native environment for membrane proteins. Despite the broad potential of charged nanodiscs, a systematic study of the influence of charged lipids on the nanodisc structure and stability has not yet been accomplished. In this paper, binary systems of zwitterionic DMPC mixed with the anionic lipids DMPG or DMPA or with the cationic synthetic DMTAP are used to prepare negatively and positively charged nanodiscs, respectively. Size exclusion chromatography analysis shows that nanodiscs can be prepared with high yield at all compositions of DMPC and DMPG, while mixtures of DMPC with either DMPA or DMTAP impair nanodisc formation. The presence of DMPG improves the stability of the nanodisc, both thermally and over time upon storage at −20 °C, as compared to pure DMPC nanodiscs. This stabilization is attributed to favourable electrostatic interactions between the anionic head of DMPG and cationic charges of the belt protein and inter-nanodisc repulsion that prevents aggregation of nanodiscs. In contrast, even small fractions of DMPA result in a faster degradation at −20 °C. These results suggest that the mixing of DMPC and DMPG provides nanodiscs that are better suited for studies of the function and structure of membrane proteins not only due to their inherent charge but also due to their improved thermal and storage stability compared to pure DMPC nanodiscs.