AbstractCarbon nanodots (CNDs) doped with Tb ions were synthesized using different synthetic routes: hydrothermal treatment of a solution containing carbon source (sodium dextran sulfate) and TbCl₃; mixing of CNDs and TbCl₃ solutions; freezing-induced loading of Tb and carbon-containing source into pores of CaCO₃ microparticles followed by hydrothermal treatment. Binding of Tb ions to CNDs (Tb-CND coupling) was confirmed using size-exclusion chromatography and manifested itself through a decrease of the Tb photoluminescence lifetime signal. The shortest Tb photoluminescence lifetime was observed for samples obtained by hydrothermal synthesis of CaCO₃ microparticles where Tb and carbon source were loaded into pores via the freezing-induced process. The same system displays an increase of Tb photoluminescence via energy transfer with excitation at 320–340 nm. Based on the obtained results, freezing-induced loading of cations into CNDs using porous CaCO₃ microparticles as reactors is proposed to be a versatile route for the introduction of active components into CNDs. The obtained CNDs with long-lived emission may be used for time-resolved imaging and visualization in living biological samples where time-resolved and long-lived luminescence microscopy is required.