Uptake of diterbium transferrin, a potential multi-photon-excited microscopy probe, into human leukemia K562 cells via a transferrin-receptor-mediated process

Our present study provided new evidence for diterbium transferrin (Tb2Tf) as a potential multi-photon-excited microscopy probe. It indicated that the Tb2Tf complex can be transported into human leukemia K562 cells via a process mediated by transferrin (Tf) receptors as an intact entity and with no obvious cellular toxicity. The supporting evidence includes the following. First, the transport kinetic behavior of Tb was compared with that of the Tf moiety. The Tb was determined by inductively coupled plasma mass spectrometry and Tf was determined by fluorescence activated cell sorting analysis. The kinetic synchronization of internalization of both Tb and Tf into human leukemia K562 cells demonstrated the Tb2Tf complex was transported into cells as a whole. Second, using confocal laser scanning microscopy, we observed the localization of Tb2Tf in the cell. This showed that the internalized Tb2Tf was mostly situated in the same perinuclear region as diferric transferrin (Fe2Tf). In addition, pretreatment with pronase largely abolished the transport process of Tb2Tf. The relative fluorescence intensities representing the uptake of Tf into the cells decreased to about 16% and the cytosolic Tb content decreased to almost the same percentage as for Tf. Furthermore, the addition of Fe2Tf can effectively inhibit transport of Tb2Tf into K562 cells. Third, no significant decrease of cell viability was observed in the presence of Tb2Tf even for 24 or 48 h by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Moreover, prospects for the use of Tb2Tf as a multi-photon-excited microscopy probe in a living system are discussed.