Research on graphene-based nanomaterials has experienced exponential growth in the last few decades, driven by their unique properties and their future potential impact on our everyday life. With the increasing production and commercialization of these materials, there is significant interest in understanding their fate in vivo. Herein, we investigated the distribution of 14C-few-layer graphene (14C-FLG) flakes (lat. dim. ∼ 500 nm) in mice over a period of one year. Furthermore, we compared the effects of repeated low-dose and acute high-dose exposure by tracheal administration. The results showed that most of the radioactivity was found in the lungs in both cases, with longer elimination times in the case of acute high-dose administration. In order to gain deeper insights into the distribution pattern, we conducted ex vivo investigations using μ-autoradiography on tissue sections, revealing the heterogeneous distribution of the material following administration. For the first time, μ-autoradiography was used to conduct a comprehensive investigation into the distribution and potential presence of FLG within lung cells isolated from the exposed lungs. The presence of radioactivity in lung cells strongly suggests internalization of the 14C-FLG particles. Overall these results show the long-term accumulation of the material in the lungs over one year, regardless of the administration protocol, and the higher biopersistence of FLG in the case of an acute exposure. These findings highlight the importance of the exposure scenario in the context of intratracheal administration, which is of interest in the evaluation of the potential health risks of graphene-based nanomaterials.