The biodistribution of extracellular vesicles (EVs) is a fundamental question in the field of circulating biomarkers, which has recently gained attention. Despite the capabilities of nuclear imaging methods such as single photon emission computed tomography (SPECT), radioisotope labeling of EVs and the use of the aforementioned methods for in vivo studies hardly can be found in the literature. In this paper we describe a novel method for the radioisotope labeling of erythrocyte-derived EVs using the 99m Tctricarbonyl complex. Moreover, the capability of the developed labeling method for in vivo biodistribution studies is demonstrated in a mouse model. We found that the intravenously administered, According to our data, only a minor fraction of the radioactive label became detached from the EVs.

The in vivo biodistribution of liposomal formulations greatly influences the pharmacokinetics of these novel drugs; therefore the radioisotope labeling of liposomes and the use of nuclear imaging methods for in vivo studies are of great interest. In the present work, a new procedure for the surface labeling of liposomes is presented using the novel 99mTc-tricarbonyl complex. Liposomes mimicking the composition of two FDA approved liposomal drugs were used. In the first step of the labeling, thiol-groups were formed on the surface of the liposomes using Traut's reagent, which were subsequently used to bind 99mTc-tricarbonyl complex to the liposomal surface. The labeling efficiency determined by size exclusion chromatography was 95%, and the stability of the labeled liposomes in bovine serum was found to be 94% over 2 hours. The obtained specific activity was 50 MBq per 1 μmol lipid which falls among the highest values reported for 99mTc labeling of liposomes. Quantitative in vivo SPECT/CT biodistribution studies revealed distinct differences between the labeled liposomes and the free 99mTc-tricarbonyl, which indicates the in vivo stability of the labeling. As the studied liposomes were non-PEGylated, fast clearance from the blood vessels and high uptake in the liver and spleen were observed.