MAGfect: a novel liposome formulation for MRI labelling and visualization of cells

Cellular entry of imaging probes, such as contrast agents for magnetic resonance imaging (MRI), is a key requirement for many molecular imaging studies, particularly imaging intracellular events and cell tracking. Here, we describe the successful development and in vitro analysis of MAGfect, a novel liposome formulation containing a lipidic gadolinium contrast agent for MRI, Gd-DOTA-Chol , designed to enter and label cells. Liposome formulation and cell incubation time were optimised for maximum cellular uptake of the imaging probe in a variety of cell lines. MRI analysis of cells incubated with MAGfect showed them to be highly MRI active. This formulation was examined further for cytotoxicity, cell viability and mechanism of cell labelling. One of the key advantages of using MAGfect as a labelling vehicle arises from its potential for additional functions, such as concomitant drug or gene delivery and fluorescent labelling. The gadolinium liposome was found to be an effective vehicle for transport of plasmid DNA (pDNA) into cells and expression levels were comparable to the commercial transfection agent Trojene.     

Proteomic analysis of Pseudomonas aeruginosa grown under magnesium limitation

In this study, large-scale qualitative and quantitative proteomic technology was applied to the analysis of the opportunistic bacterial pathogen Pseudomonas aeruginosa grown under magnesium limitation, an environmental condition previously shown to induce expression of various virulence factors. For quantitative analysis, whole cell and membrane proteins were differentially labeled with isotope-coded affinity tag (ICAT) reagents and ICAT reagent-labeled peptides were separated by two-dimensional chromatography prior to analysis by electrospray ionization-tandem mass spectrometry (ESI-MS/MS) in an ion trap mass spectrometer (ITMS). To increase the number of protein identifications, gas-phase fractionation (GPF) in the m/z dimension was employed for analysis of ICAT peptides derived from whole cell extracts. The experiments confirmed expression of 1331 P. aeruginosa proteins of which 145 were differentially expressed upon limitation of magnesium. A number of conserved Gram-negative magnesium stress-response proteins involved in bacterial virulence were among the most abundant proteins induced in low magnesium. Comparative ICAT analysis of membrane versus whole cell protein indicated that growth of P. aeruginosa in low magnesium resulted in altered subcellular compartmentalization of large enzyme complexes such as ribosomes. This result was confirmed by 2-D PAGE analysis of P. aeruginosa outer membrane proteins. This study shows that large-scale quantitative proteomic technology can be successfully applied to the analysis of whole bacteria and to the discovery of functionally relevant biologic phenotypes.     

Rapid determination of moisture and fat in meats by microwave and nuclear magnetic resonance analysis--PVM 1:2003

A peer-verified method is presented for the determination of percent moisture and fat in meat products by microwave drying and nuclear magnetic resonance (NMR) analysis. The method involves determining the moisture content of meat samples by microwave drying and using the dried sample to determine the fat content by NMR analysis. Both the submitting and peer laboratories analyzed 5 meat products by using the CEM SMART system (moisture) and the SMART Trac (fat). The samples, which represented a range of products that meat processors deal with daily in plant operations, included the following: (1) fresh ground beef, high-fat; (2) deboned chicken with skin; (3) fresh pork, low-fat; (4) all-beef hot dogs; and (5) National Institute of Standards and Technology Standard Reference Material. The results were compared with moisture and fat values derived from AOAC-approved methods, 950.46 (Forced Air Oven Drying) and 960.39 (Soxhlet Ether Extraction).     

TMTHSI,a superior 7-membered ring alkyne containing reagent for strain-promoted azide–alkyne cycloaddition reactions

We describe the development of TMTH-SulfoxImine (TMTHSI) as a superior click reagent. This reagent combines a great reactivity, with small size and low hydrophobicity and compares outstandingly with existing click reagents. TMTHSI can be conveniently functionalized with a variety of linkers allowing attachment of a diversity of small molecules and (peptide, nucleic acid) biologics.

TMTHSI was developed as new reagent for strain-promoted azide–alkyne cycloaddition reactions, enabling connection of a diversity of small to large molecular constructs.     

Coating MWNTs with Cu2O of different morphology by a polyol process

Homogeneous cuprous oxide (Cu₂O) nanoparticles with size of 8-10 nm are deposited on multiwall carbon nanotubes (MWNTs) by a polyol process using Cu(CH₃COO)₂·H₂O as a precursor and diethylene glycol as both solvent and reducing agent. The composition of the resulting Cu₂O/MWNTs composites is confirmed by XRD pattern, XPS spectrum and HRTEM images. With the change of the reaction conditions, it is found that Cu₂O nanoparticles on the surface of MWNTs can be leafage-like or big spherical particle coated on the surface of MWNTs. HRTEM images indicate that all the leafage-like and big spherical particles are assembled by small Cu₂O particles with size of about 2-5 nm. With the assistance of FTIR spectrum, a tentative mechanism is proposed for the formation of Cu₂O nanoparticles with different morphologies on the surface of MWNTs.     

Linking reactions of living polymers with bromomethylbenzene derivatives: Synthesis and characterization of star homopolymers and graft copolymers with polyelectrolyte branches

Anionic polymerization techniques utilizing 1,2,4,5-tetra(bromomethyl)- benzene as the linking agent were employed for the synthesis of four-arm star polymers with poly(tert-butyl methacrylate) (PtBuMA), poly(methyl methacrylate), poly(tert-butylacrylate) (PtBuA), or poly(2-vinylpyridine) (P2VP) branches. This work was extended through the “grafting onto” method, in combination with anionic polymerization techniques, to synthesize graft copolymers consisting of polystyrene backbones and PtBuA, PtBuMA, or P2VP branches. Postpolymerization reactions were performed to produce graft copolymers with polyelectrolyte branches. Crosslinking reactions were observed in some of the graft materials several months after their preparation. © 1999 John Wiley & Sons, Inc. J Polym Sci A: Polym Chem 37: 4337–4350, 1999     

Synthesis and characterization of multiarm star-branched polyisobutylenes: Effect of arm molecular weight

A series of star-branched polyisobutylenes with varying arm molecular weights was synthesized using the 2-chloro-2,4,4-trimethylpentane/TiCl₄/pyridine initiating system and divinylbenzene (DVB) as a core-forming comonomer (linking agent). The resulting star-branched polymers were characterized with regard to the weight-average number of arms per star molecule (N̄_w) and dilute solution viscosity behavior. As the molecular weight of the arm (M̄_{w, arm}) was increased, dramatically longer star-forming reaction times were needed to produce fully developed star polymers. It was calculated that N̄_w varied from 50 to 5 as the M̄_{w, arm} was increased from 13,000 to 54,000 g/mol. The radius of gyration, R_g, of the star polymers was observed to increase as M̄_{w, arm} was increased. The solution properties of the star polymers were evaluated in heptane using dilute solution viscometry. It was determined that the stars had a much higher [η] compared to the respective linear PIB arms, but a much lower [η] compared to a hypothetical linear analog of an equivalent molecular weight. The dependence of [η] on temperature for the stars and linear arms was very small over the temperature range 25 to 75°C, with only a very slight decrease with increasing temperature. [η]_star was also determined to increase with increasing M̄_{w, arm}, but decrease with increasing M̄_{w, star}. The branching coefficient, g′, calculated for the stars at 25°C, increased as N̄_w decreased and agre ed well with literature values for other star polymer systems. © 1997 John Wiley & Sons, Inc. J Polym Sci A: Polym Chem 35: 3767–3778, 1997     

A new synthesis of aryl substituted quinazolin-4(1H)-ones

Treatment of a variety of substituted 2-aminobenzonitriles with formic acid under strong acid catalysis provides the corresponding quinazolin-4(1H)-ones in good yield. A potential reaction pathway is described.     

Dilute solution properties of asymmetric six-arm star polystyrenes

Six-arm star polystyrenes having varying numbers of short and long arms attached to the same molecule have been synthesized by anionic polymerization. The molecules have been characterized by high resolution size exclusion chromatography using multiangle light scattering and viscosity detectors. This technique has allowed the radii of gyration and intrinsic viscosities to be measured for stars with each possible combination of arms. The branching parameters g and g′ are computed and compared with theoretical expectations. It is found that short arms add preferentially to the stars, because of reduced steric effects. The molecule with one long and five short arms exhibits behavior closest to that of a linear chain (largest branching ratios). The effect of arm polydispersity on solution properties of stars is discussed. © 1995 John Wiley & Sons, Inc.