We report the development of functionalized superparamagnetic iron oxide nanoparticles with a PEG-modified, phospholipid micelle coating, and their delivery into living cells. The size of the coated particles, as determined by dynamic light scattering and electron microscopy, was found to be between 12 and 14 nm. The PEG-phospholipid coating resulted in high water solubility and stability, and the functional groups of modified PEG allowed for bioconjugation of various moieties, including a fluorescent dye and the Tat peptide. Efficient delivery of the functionalized nanoparticles into living cells was confirmed by fluorescence microscopy, relaxation time measurements, and magnetic resonance imaging (MRI). This demonstrates the feasibility of using functionalized magnetic nanoparticles with uniform (approximately 10 nm) sizes as an MRI contrast agent for intracellular molecular imaging in deep tissue. These micelle-coated iron oxide nanoparticles offer a versatile platform for conjugation of a variety of moieties, and their small size confers advantages for intracellular molecular imaging with minimal perturbation.

Purpose:To evaluate the effect of coating thickness on the relaxivity of iron oxide nanoparticles. Materials and Methods:Monocrystalline superparamagnetic iron oxide nanoparticles (MIONs), coated with a polyethylene glycol (PEG)-modified, phospholipid micelle coating, with different PEG molecular weights, were prepared. The particle diameters were measured with dynamic light scattering (DLS) and electron microscopy (EM). The R 1 and R 2 of MIONs were measured using a bench-top nuclear magnetic resonance (NMR) relaxometer. pH was varied for some measurements. Monte Carlo simulations of proton movement in a field with nanometer-sized magnetic inhomogeneities were performed. Results:Increasing the molecular weight of the PEG portion of the micelle coating increased overall particle diameter. As coating thickness increases, the R 2 decreases and the R 1 increases. Changing pH has no effect on relaxivity. The Monte Carlo simulations suggest that the effect of coating size on R 2 relaxivity is determined by two competing factors: the physical exclusion of protons from the magnetic field and the residence time for protons within the coating zone. Conclusion:Coating thickness can significantly impact the R 2 , and the R 2 /R 1 ratio, of a MION contrast agent. An understanding of the relationship between coating properties and changes in relaxivity is critical for designing magnetic nanoparticle probes for molecular imaging applications using MRI.

Magnetic resonance imaging (MRI) is routinely used to obtain anatomical images that have greatly advanced biomedical research and clinical health care today, but the full potential of MRI in providing functional, physiological, and molecular information is only beginning to emerge. In this work, we sought to provide a gene expression marker for MRI based on bacterial magnetosomes, tiny magnets produced by naturally occurring magnetotactic bacteria. Specifically, magA, a gene in magnetotactic bacteria known to be involved with iron transport, is expressed in a commonly used human cell line, 293FT, resulting in the production of magnetic, iron-oxide nanoparticles by these cells and leading to increased transverse relaxivity. MRI shows that these particles can be formed in vivo utilizing endogenous iron and can be used to visualize cells positive for magA. Synthetic superparamagnetic iron-oxide (SPIO) nanoparticles have been widely used for targeted molecular imaging applications (1-5). One major application is in vivo tracking of stem cells (6,7) and tumor progression (5). Labeling nonphagocytic cells in culture using modified particles, followed by transplantation or transfusion into living organisms, has made it possible to monitor cellular distribution in vivo, including cell migration and trafficking.A limitation of using synthetic SPIO is the need to label cells in vitro with presynthesized nanoparticles prior to cell transplant. As a result, particle concentration within cells decreases over time as the cells grow and divide, and particles cannot be readily linked directly to in vivo gene expression. One way to overcome this is to utilize a genetic approach. Green fluorescent protein (GFP) is perhaps the most well-known genetic marker for optical imaging. Magnetic resonance spectroscopy (MRS) has been used to detect creatine kinase (8) and chemical shift imaging (CSI) to observe betagalactosidase (9) activity. MRI gene expression strategies thus far include detection of beta-galactosidase activity (10,11), frequency-selective targeting of amide protons of expressed proteins (12), and expression of natural iron homeostasis proteins such as the transferrin receptor (13) and ferritin (14,15). For the transferrin receptor approach, administration of exogenous transferrin coupled to magnetic particles is required. Thus far, only ferritin exists as a purely in vivo superparamagnetic MRI marker. Although the relaxivity of ferritin is dependent on factors such as iron loading, data obtained on solutions of iron-containing materials suggest iron-oxide particles could provide higher relaxivity (16). In the present work, we report the gene-mediated cellular production of magnetic iron-oxide nanoparticles of the same composition as synthetic SPIO preparations using a gene present in magnetotactic bacteria, making it a possible MRI gene reporter.Magnetotactic bacteria, a diverse set of Gram-negative bacteria that exhibit motility thought to be directed by the earth's magnetic field (17), produce magnetosomes which are na...