The in vivo tracing of the biodistribution of extracellular vesicles (EVs) is a pre-requisite in identifying their target cells and understanding their function. Although luorescent labelling of EVs is already used, radiolabelling can provide more details in understanding biodistribution of EVs. In the present paper we report radiolabelling of bone marrow-derived EVs and in vivo tracing of their biodistribution. EVs isolated from the bone marrow supernatant of űő7ŰL/6 mice were labelled with the technetium-99m (99mTc) isotope. Labelling was eficient and labelled EVs were stable during the 2Ő hours follow-up. Detection of labelled EVs after intravenous injection in mice was performed using ex vivo measurements and in vivo imaging. Ex vivo examinations revealed that at Ő hours post-injection, the highest accumulation rate was in the liver, kidney, spleen and femur epiphysis. In vivo imaging using nanoSPEűT/űT conirmed the ex vivo examinations and demonstrated slow elimination of the radioactivity, 2Ő hours post- injection the bone marrow-containing epiphysis and lymph nodes showed the highest retention values; liver, spleen and kidney were also clearly detectable. In summary, labelling of bone marrow-derived EVs with 99mTc coupled with SPEűT/űT detection was a reliable method for quantitative distribution studies of EVs in vivo
