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Nanoparticles used for biologicaland biomedical applications encountera host of extracellular proteins. These proteins rapidly adsorb ontothe nanoparticle surface, creating a protein corona. Poly(ethyleneglycol) can reduce, but not eliminate, the nonspecific adsorptionof proteins. As a result, the adsorbed proteins, rather than the nanoparticleitself, determine the cellular receptors used for binding, the internalizationmechanism, the intracellular transport pathway, and the subsequentimmune response. Using fluorescence microscopy and flow cytometry,we first characterize a set of polystyrene nanoparticles in whichthe same adsorbed protein, bovine serum albumin, leads to bindingto two different cell surface receptors: native albumin receptorsand scavenger receptors. Using a combination of circular dichroismspectroscopy, isothermal titration calorimetry, and fluorescence spectroscopy,we demonstrate that the secondary structure of the adsorbed bovineserum albumin protein controls the cellular receptors used by theprotein–nanoparticle complexes. These results show that proteinsecondary structure is a key parameter in determining the cell surfacereceptor used by a protein–nanoparticle complex. We expectthis link between protein structure and cellular outcomes will providea molecular basis for the design of nanoparticles for use in biologicaland biomedical applications. [ABSTRACT FROM AUTHOR]

An ultrafast UV pump/IR probe study is presented to show triplet organometallic reactivity under ambient conditions.

The use of nanoparticles (NPs) in biology andmedicine requiresa molecular-level understanding of how NPs interact with cells ina physiological environment. A critical difference between well-controlled in vitroexperiments and in vivoapplicationsis the presence of a complex mixture of extracellular proteins. Ithas been established that extracellular serum proteins present inblood will adsorb onto the surface of NPs, forming a “proteincorona”. Our goal was to understand how this protein layeraffected cellular-level events, including NP binding, internalization,and transport. A combination of microscopy, which provides spatialresolution, and spectroscopy, which provides molecular information,is necessary to probe protein–NP–cell interactions.Initial experiments used a model system composed of polystyrene NPsfunctionalized with either amine or carboxylate groups to providea cationic or anionic surface, respectively. Serum proteins adsorbonto the surface of both cationic and anionic NPs, forming a net anionicprotein–NP complex. Although these protein–NP complexeshave similar diameters and effective surface charges, they show theexact opposite behavior in terms of cellular binding. In the presenceof bovine serum albumin (BSA), the cellular binding of BSA–NPcomplexes formed from cationic NPs is enhanced, whereas the cellularbinding of BSA–NP complexes formed from anionic NPs is inhibited.These trends are independent of NP diameter or cell type. Similarresults were obtained for anionic quantum dots and colloidal goldnanospheres. Using competition assays, we determined that BSA–NPcomplexes formed from anionic NPs bind to albumin receptors on thecell surface. BSA–NP complexes formed from cationic NPs areredirected to scavenger receptors. The observation that similar NPswith identical protein corona compositions bind to different cellularreceptors suggested that a difference in the structure of the adsorbedprotein may be responsible for the differences in cellular bindingof the protein–NP complexes. Circular dichroism spectroscopy,isothermal titration calorimetry, and fluorescence spectroscopy showthat the structure of BSA is altered following incubation with cationicNPs, but not anionic NPs. Single-particle-tracking fluorescence microscopywas used to follow the cellular internalization and transport of protein–NPcomplexes. The single particle-tracking experiments show that theprotein corona remains bound to the NP throughout endocytic uptakeand transport. The interaction of protein–NP complexes withcells is a challenging question, as the adsorbed protein corona controlsthe interaction of the NP with the cell; however, the NP itself altersthe structure of the adsorbed protein. A combination of microscopyand spectroscopy is necessary to understand this complex interaction,enabling the rational design of NPs for biological and medical applications. [ABSTRACT FROM AUTHOR]

Nanoparticles are increasingly important for biologicalapplicationsranging from drug delivery to cellular imaging. In the course of theseapplications, nanoparticles are exposed to a complex environment ofextracellular proteins that can be adsorbed onto the surface of thenanoparticle, altering nanoparticle–cell interactions. We haveinvestigated how proteins found in blood serum affect the bindingof nanoparticles to the surface of cells. Using fluorescence microscopy,we find that the cellular binding of cationic nanoparticles is enhancedby the presence of serum proteins, while the binding of anionic nanoparticlesis inhibited. We have determined that this difference in cellularbinding is due to the use of distinct cellular receptors. Competitionassays, quantified with flow cytometry, show that the protein–nanoparticlecomplex formed from the cationic nanoparticles binds to scavengerreceptors on the cell surface. Interestingly, the protein–nanoparticlecomplex formed from anionic nanoparticles binds to native proteinreceptors. As nanoparticles become increasingly important for in vivoapplications, we expect these results will informthe design of nanoparticles with improved cellular binding. [ABSTRACT FROM AUTHOR]

Abstract: Endocytosis, the internalization and transport of extracellular cargo, is an essential cellular process. The ultimate step in endocytosis is the intracellular degradation of extracellular cargo for use by the cell. While live cell imaging and single particle tracking have been well-utilized to study the internalization and transport of cargo, the final degradation step has required separate biochemical assays. We describe the use of self-quenched endocytic cargo to image the intracellular transport and degradation of endocytic cargo directly in live cells. We first outline the fluorescent labeling and quantification of two common endocytic cargos: a protein, bovine serum albumin, and a lipid nanoparticle, low-density lipoprotein. In vitro measurements confirm that self-quenching is a function of the number of fluorophores bound to the protein or particle and that recovery of the fluorescent signal occurs in response to enzymatic degradation. We then use confocal fluorescence microscopy and flow cytometry to demonstrate the use of self-quenched bovine serum albumin with standard fluorescence techniques. Using live cell imaging and single particle tracking, we find that the degradation of bovine serum albumin occurs in an endo-lysosomal vesicle that is positive for LAMP1. [Copyright &y& Elsevier]

The article offers information on the achievements and performance of the Elgar Society from January 1 to December 31, 2008. The main topics of the discussion during the 2008 Annual General Meeting include the questionnaire regarding the membership's expectations, a new scheme for the Society, and new recordings and other initiatives. It notes that the web site of the society continues to offer the general public with a detailed reference source regarding the life and music of Edward Elgar.

Gene delivery offers the promise of treatment for a range of human diseases. Although carried out initially with modified viruses, the use of synthetic molecules, including polymers, lipids and peptides, has extended the possibilities greatly for rationally designed vectors tailored to individual gene-delivery applications. Underlying the rational design of gene-delivery vectors is the need to understand the individual steps of the gene-delivery pathway. Using new methods in fluorescence microscopy, it is now possible to isolate individual steps along the gene-delivery pathway to characterize the mechanisms of cellular binding, cellular internalization and nuclear entry. This review describes the advances made in the gene-delivery field with the assistance of fluorescence microscopy. The focus of this review is the use of synthetic gene-delivery vectors, especially polyethylenimine, and the live-cell imaging and single-particle tracking techniques that reveal the intracellular dynamics of the gene-delivery process. [ABSTRACT FROM AUTHOR]