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In vivo cell tracking is vital for understanding migrating cell populations, particularly cancer and immune cells. Magnetic resonance (MR) imaging for long-term tracking of transplanted cells in live organisms requires cells to effectively internalize Gd(III) contrast agents (CAs). Clinical Gd(III)-based CAs require high dosing concentrations and extended incubation times for cellular internalization. To combat this, we have devised a series of Gd(III)-gold nanoconjugates (Gd@AuNPs) with varied chelate structure and nanoparticle-chelate linker length, with the goal of labeling and imaging breast cancer cells. These new Gd@AuNPs demonstrate significantly enhanced labeling compared to previous Gd(III)-gold-DNA nanoconstructs. Variations in Gd(III) loading, surface packing, and cell uptake were observed among four different Gd@AuNP formulations suggesting that linker length and surface charge play an important role in cell labeling. The best performing Gd@AuNPs afforded 23.6 ± 3.6 fmol of Gd(III) per cell at an incubation concentration of 27.5 μM-this efficiency of Gd(III) payload delivery (Gd(III)/cell normalized to dose) exceeds that of previous Gd(III)-Au conjugates and most other Gd(III)-nanoparticle formulations. Further, Gd@AuNPs were well-tolerated in vivo in terms of biodistribution and clearance, and supports future cell tracking applications in whole-animal models., Competing Interests: Notes The authors declare no competing financial interest.

Calcium [Ca(II)] is a fundamental transducer of electrical activity in the central nervous system (CNS). Influx of Ca(II) into the cytosol is responsible for action potential initiation and propagation, and initiates interneuronal communication via release of neurotransmitters and activation of gene expression. Despite the importance of Ca(II) in physiology, it remains a challenge to visualize Ca(II) flux in the central nervous system (CNS) in vivo. To address these challenges, we have developed a new generation, Ca(II)-activated MRI contrast agent that utilizes ethyl esters to increase cell labeling and prevent extracellular divalent Ca(II) binding. Following labeling, the ethyl esters can be cleaved, thus allowing the agent to bind Ca(II), increasing relaxivity and resulting in enhanced positive MR image contrast. The ability of this probe to discriminate between extra- and intracellular Ca(II) may allow for spatiotemporal in vivo imaging of Ca(II) flux during seizures or ischemia where large Ca(II) fluxes (1-10 μM) can result in cell death.

Multiple imaging modalities are often required for in vivo imaging applications that require both high probe sensitivity and excellent spatial and temporal resolution. In particular, MR and optical imaging are an attractive combination that can be used to determine both molecular and anatomical information. Herein, we describe the synthesis and in vivo testing of two multimeric NIR-MR contrast agents that contain three Gd(III) chelates and an IR-783 dye moiety. One agent contains a PEG linker and the other a short alkyl linker. These agents label cells with extraordinary efficacy and can be detected in vivo using both imaging modalities. Biodistribution of the PEGylated agent shows observable fluorescence in xenograft MCF7 tumors and renal clearance by MR imaging.

Nanodiscs are monodisperse, self-assembled discoidal particles that consist of a lipid bilayer encircled by membrane scaffold proteins (MSP). Nanodiscs have been used to solubilize membrane proteins for structural and functional studies and deliver therapeutic phospholipids. Herein, we report on tetramethylrhodamine (TMR) tagged nanodiscs that solubilize lipophilic MR contrast agents for generation of multimodal nanoparticles for cellular imaging. We incorporate both multimeric and monomeric Gd(III)-based contrast agents into nanodiscs and show that particles containing the monomeric agent (ND2) label cells with high efficiency and generate significant image contrast at 7 T compared to nanodiscs containing the multimeric agent (ND1) and Prohance, a clinically approved contrast agent.

We describe the design, synthesis and in vitro evaluation of a multimodal and multimeric contrast agent. The agent consists of three macrocyclic Gd(III) chelates conjugated to a fluorophore and possesses high relaxivity, water solubility, and is nontoxic. The modular synthesis is amenable for the incorporation of a variety of fluorophores to generate molecular constructs for a number of applications.

Cell tracking in vivo with MR imaging requires the development of contrast agents with increased sensitivity that effectively label and are retained by cells. Most clinically approved Gd(III)-based contrast agents require high incubation concentrations and prolonged incubation times for cellular internalization. Strategies to increase contrast agent permeability have included conjugating Gd(III) complexes to cell penetrating peptides, nanoparticles, and small molecules which have greatly improved cell labeling but have not resulted in improved cellular retention. To overcome these challenges, we have synthesized a series of lipophilic Gd(III)-based MR contrast agents that label cell membranes in vitro. Two of the agents were synthesized with a multiplexing strategy to contain three Gd(III) chelates (1 and 2) while the third contains a single Gd(III) chelate (3). These new agents exhibit significantly enhanced labeling and retention in HeLa and MDA-MB-231-mcherry cells compared to agents that are internalized by cells (4 and Prohance).

LnDOTA-tetraamide chelates (DOTA=1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) have received considerable recent attention as a result of their potential to act as PARACEST contrast agents for magnetic resonance imaging (MRI). Although PARACEST agents afford several advantages over conventional contrast agents they suffer from substantially higher detection limits; thus, improving the effectiveness of LnDOTA-tetraamide chelates is an important goal. In this study we investigate the potential to extend conformational control of LnDOTA-type ligands to those applicable to PARACEST. Furthermore, the question of whether δ- rather than α-substitution of the pendant arms could be used to control the chelate coordination geometry is addressed. Although δ-substitution does influence coordination geometry it does not afford control. However, it can play an important role in governing the conformation of the amide substituent relative to the chelate in such as way that suggests a PARACEST agent could be designed that has detection limits at least as low as a conventional MRI contrast agent., (Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)