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Cryptophanes have been shown to be a promising tool in xenon magnetic resonance imaging (MRI) applications. The affinity of xenon for molecular host structures such as cryptophane-A (CrA) and the large xenon chemical shift that can be detected in the presence of such hosts has motivated the synthesis of a number of cryptophane-based MRI biosensors. Bifunctional biosensors that incorporate both a fluorophore and a cryptophane moiety are flexible tools for evaluating the intracellular localization and quantifying cellular uptake/binding of such xenon hosts in conjunction with testing their MRI capabilities. In this study, we compare the performance of a new compound that bears tetramethylrhodamine (TAMRA), CrA-PEG-TAMRA (PEG=polyethylene glycol), with our previously synthesized compound in which fluorescein (FAM) was the optical reporter. We demonstrate that these biosensors are suitable candidates for MRI-based cell-tracking studies; specifically, the new addition of a rhodamine derivative will allow for multiplexing experiments and showed slightly improved NMR spectroscopic behavior. Importantly, both biosensors display equally good cell-labeling efficiency, which facilitates the acquisition of high-quality xenon magnetic resonance images in two different cell lines. This study describes a simple synthetic approach for the production of biologically compatible bifunctional compounds. These biosensors can be used as they are for cell labeling but can also serve as building blocks for further functionalization and cryptophane-based xenon biosensor design with MRI and fluorescence multiplexing options.

Riboflavin is an essential vitamin for cellular metabolism and is highly upregulated in metabolically active cells. Consequently, targeting the riboflavin carrier protein (RCP) may be a promising strategy for labeling cancer and activated endothelial cells. Therefore, Ultrasmall SuperParamagnetic Iron Oxide nanoparticles (USPIO) were adsorptively coated with the endogenous RCP ligand flavin mononucleotide (FMN), which renders them target-specific and fluorescent. The core diameter, surface morphology and surface coverage of the resulting FMN-coated USPIO (FLUSPIO) were evaluated using a variety of physico-chemical characterization techniques (TEM, DLS, MRI and fluorescence spectroscopy). The biocompatibility of FLUSPIO was confirmed using three different cell viability assays (Trypan blue staining, 7-AAD staining and TUNEL). In vitro evaluation of FLUSPIO using MRI and fluorescence microscopy demonstrated high labeling efficiency of cancer cells (PC-3, DU-145, LnCap) and activated endothelial cells (HUVEC). Competition experiments (using MRI and ICP-MS) with a 10- and 100-fold excess of free FMN confirmed RCP-specific uptake of the FLUSPIO by PC-3 cells and HUVEC. Hence, RCP-targeting via FMN may be an elegant way to render nanoparticles fluorescent and to increase the labeling efficacy of cancer and activated endothelial cells. This was shown for FLUSPIO, which due to their high T2-relaxivity, are favorably suited for MR cell tracking experiments and cancer detection in vivo.

Nano research 9(5), 1319-1333 (2016). doi:10.1007/s12274-016-1028-7, Published by Tsinghua Press, [S.l.]