Electrolyte‐gated organic field‐effect transistors (EGOFETs) are subject to intense research for biosensing in fluids. The ability of these devices to quantify a variety of chemical and biological molecules sensitively and selectively, but ex situ, has been widely demonstrated. However, continuous monitoring of analyte‐receptor interactions in real time by EGOFETs, in high demand for practical applications, has rarely been explored. Here, an EGOFET array integrated with a microfluidic device, for real‐time detection of the hybridization of DNA is presented. The integrated devices exhibit highly reproducible electrical performance in the array and can operate under different electrical stresses over > 1 h with 85–96% figures of merit retention. The utility of the devices is demonstrated to detect the hybridization of a complementary target DNA in 10 × 10 −3 m phosphate‐buffered saline (1 × PBS) selectively with a temporal resolution of < 1 s in a flow of analyte with no incubation step. The detection time is < 30 s and the relative standard deviation of the sensing reproducibility is < 15% under the target concentration of 100 × 10 −9 m .

The rapid detection of disease specific biomarkers in a clinically relevant range using a low-cost sensor can facilitate the development of individual treatment plans for a given patient, known as precision, personalized or genomic medicine. In the recent decade Electrolyte-Gated Organic Field Effect Transistors (EGOFETs), a subtype of OFETs where the dielectric is replaced by an electrolyte, have attracted a great deal of attention for sensing applications. This is due to their capacity to operate at low voltage (< 1 volt) in physiological like media. Although EGOFET based biosensors have been shown to specifically detect biomolecules with high sensitivity and selectivity; the stability, reproducibility, and performance required to reach the desired market are not yet achieved. In this contribution, we describe the development of a stable and reproducible EGOFET sensor that is able to detect biomolecules selectively in real-time. Facile and scalable techniques are used to prepare arrays of these devices. The selectivity of individual EGOFETs is investigated by immobilization of specific ligands to the target molecule of interest on the gate electrode within a microfluidic flow cell.

The development of synthetic strategies to control the molecular organization (and inherently linked optoelectronic properties) of conjugated polymers is critical for the development of efficient light-emitting devices. Here, we report a facile route using sol gel chemistry to promote the formation of the beta-phase through the covalent -grafting of poly[(9,9dioctylfluorene)-co-(9,9-bis(8-hydroxyoctyl)fluorene)] (PFO-OH) to poly(oxyalkylene)/siloxane hybrids known as ureasils, due to the urea linkages binding the organic and inorganic components. Although grafting occurs within the siliceous domains, the degree of branching of the organic backbone determines the packing of the PFO-OH chains within the ureasil framework. Moreover, photoluminescence studies indicate that physical confinement also plays a key role in promoting the evolution of the beta-phase of PFO-OH as the sol gel transition proceeds. Spectroscopic and structural analyses reveal that dibranched ureasils promote linear packing of the PFO-OH chains, while tribranched ureasils exhibit a more open, distorted structure that restricts the packing efficacy and reduces the number of covalent anchorages. These results indicate that the organic inorganic hybrid structure induces distinct levels of beta-phase formation and that covalent grafting is a versatile approach to design novel poly(fluorene) hybrid materials with tailored optical properties.

Suzuki cross-coupling polymerisation of aryldibromides and aryldiboronate esters in a sodium dodecyl sulfate (SDS)-stabilised miniemulsion provides a versatile and direct route to fluorescent conjugated polymer nanoparticles (CPNs). These nanoparticles have a conjugated backbone based on poly(9,9-dioctylfluorene) (PFO), however, significant structural diversity is introduced by incorporation of electron withdrawing, heterocyclic comonomers (5–50mol.%) in order to tune the emission wavelengths from blue to far-red/near-infrared. The robust nature of the polymerisation methodology allows for rapid assessment of the relationship between polymer composition, chain morphology and optical properties of the resultant CPNs. Moreover, the CPNs (after a simple and rapid purification step) can be used directly in fluorescence-based intracellular labelling experiments (in HCT116 cells), in which they display low cytotoxicity at biologically-useful concentrations.

The present study involves an investigation of the formation, characterisation and triggered-degradation of mixed dispersions involving cationic poly(vinylamine-co-bis(ethyl vinylamine) ether) (PVAM-BEV-AME) microgel (MG) particles and partially oxidised dextran (Dexox). In this approach to colloidal hydrogel composite formation, imine bonds were formed by reaction between aldehyde groups of Dexox and the primary amine groups on the MG particles. The composite hydrogels contained MG particles that were externally cross-linked by Dexox to form an elastically effective network with high storage modulus (G') values and low tan delta (=G"/G', where G" is the loss modulus) values. The G' values for the MG Dexox gels increased exponentially with increasing mass ratio (MR) of Dexox to MG. Interestingly, the yield strains determined from rheology also increased with MR and yield strains of up to 130% were measured. Au nanoparticles of comparable size to the Dexox chains adsorbed to the surface of the MG particles, which suggests that the pore size of the MG particles may have been smaller than that of the Dexox coils. The MG Dexox gels were also subjected to acidic conditions to demonstrate pH-triggered gel network breakdown via imine bond cleavage. We show that new PVAM MG/aldehyde mixtures studied here for the first time form ductile and versatile colloidal gels and our new method provides a route to increasing ductility of hydrogels containing MG particles. (C) 2014 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY license.

Discovering the function of an unknown protein, particularly one with neither structural nor functional correlates, is a daunting task. Interaction analyses determine binding partners, whereas DNA transfection, either transient or stable, leads to intracellular expression, though not necessarily at physiologically relevant levels. In theory, direct intracellular protein delivery (protein transduction) provides a conceptually simpler alternative, but in practice the approach is problematic. Domains such as HIV TAT protein are valuable, but their effectiveness is protein specific. Similarly, the delivery of intact proteins via endocytic pathways (e.g. using liposomes) is problematic for functional analysis because of the potential for protein degradation in the endosomes/lysosomes. Consequently, recent reports that microspheres can deliver bio-cargoes into cells via a non-endocytic, energy-independent pathway offer an exciting and promising alternative for in vitro delivery of functional protein. In order for such promise to be fully exploited, microspheres are required that (i) are stably linked to proteins, (ii) can deliver those proteins with good efficiency, (iii) release functional protein once inside the cells, and (iv) permit concomitant tracking. Herein, we report the application of microspheres to successfully address all of these criteria simultaneously, for the first time. After cellular uptake, protein release was autocatalyzed by the reducing cytoplasmic environment. Outside of cells, the covalent microsphere-protein linkage was stable for ≥90 h at 37 °C. Using conservative methods of estimation, 74.3% ± 5.6% of cells were shown to take up these microspheres after 24 h of incubation, with the whole process of delivery and intracellular protein release occurring within 36 h. Intended for in vitro functional protein research, this approach will enable study of the consequences of protein delivery at physiologically relevant levels, without recourse to nucleic acids, and offers a useful alternative to commercial protein transfection reagents such as Chariot™. We also provide clear immunostaining evidence to resolve residual controversy surrounding FACS-based assessment of microsphere uptake.

Poly(p-phenylenevinylene)s (PPVs) with single reactive end groups have been prepared with high molecular weights, narrow polydispersities (ĐM) and excellent end functionality (f). PPVs functionalised with α-bromoester end groups are effective macroinitiators in the atom transfer radical polymerisation (ATRP) of methyl methacrylate (MMA).

A range of stable emulsions of spherical and rod-like conjugated polymer nanoparticles (CPN) were synthesized via Suzuki-Miyaura cross-coupling reactions of 9,9-dioctylfluorene-2,7-diboronic acid bis(1,3-propanediol) ester with a number of different dibromoarene monomers in xylene, stabilized in water by the nonionic surfactant, Triton X-102. High molar mass poly(9,9-dioctylfluorene) (PF8), poly(9,9-dioctylfluorene-alt-benzothiadiazole) (PF8BT), poly(9,9-dioctylfluorene-alt-4-sec-butylphenyldiphenylamine) (PF8TAA) and poly(9,9-dioctylfluorene-alt-bithiophene) (PF8T2) emulsions were obtained, at high overall conjugated polymer concentrations (up to 11,000 ppm), in the presence of the palladium complex, (IPr*)PdCl2(TEA) and base, tetraethylammonium hydroxide, in nitrogen atmosphere at 30 degrees C after 24-48 h. TEM analysis of the PF8 and PF8T2 emulsions revealed regular rod-like structures, up to 200 nm in length with aspect ratios of 4-5. PF8BT and PF8TAA formed spherical particles with diameters of between 20-40 nm in TEM analysis. UV-vis absorption spectra of the PF8 emulsions indicated high levels of ordered beta-phase configuration (9-10%) in their respective nanoparticles. Absolute photoluminescence quantum yields (Phi) of 21-25% were recorded for these emulsions.

Analysis of protein function in a cellular context ideally requires physiologically representative levels of that protein. Thus conventional nucleic acid-based transfection methods are far from ideal owing to the over expression that generally results. Likewise, fusions with protein transduction domains can be problematic whilst delivery via liposomes/nanoparticles typically results in endosomal localisation. Recently, polymer microspheres have been reported to be highly effective at delivering proteins into cells and thus provide a viable new alternative for protein delivery (protein transduction). Herein we describe the successful delivery of active ribonuclease A into HeLa cells via novel polymer core-silica shell microspheres. Specifically, poly(styrene-co-vinylbenzylisothiouronium chloride) core particles, generated by dispersion polymerisation, were coated with a poly(styrene-co-trimethoxysilylpropyl methacrylate) shell. The resultant core-shell morphology was characterised by transmission electron, scanning electron and confocal fluorescence microscopies, whilst size and surface charge was assessed by dynamic light scattering and zeta-potential measurements, respectively. Subsequently, ribonuclease A was coupled to the microspheres using simple carbodiimide chemistry. Gel electrophoresis confirmed and quantified the activity of the immobilised enzyme against purified HeLa RNA. Finally, the polymer-protein particles were evaluated as protein-transduction vectors in vitro to deliver active ribonuclease A to HeLa cells. Cellular uptake of the microspheres was successful and resulted in reduced levels of both intracellular RNA and cell viability.

The immobilization of functional biomolecules to surfaces is a critical process for the development of biosensors for disease diagnostics. In this work we report the patterned attachment of single chain fragment variable (scFv) antibodies to the surface of metal oxides by the photodeprotection of self-assembled monolayers, using near-UV light. The photodeprotection step alters the functionality at the surface; revealing amino groups that are utilized to bind biomolecules in the exposed regions of the substrate only. The patterned antibodies are used for the detection of specific disease biomarker proteins in buffer and in complex samples such as human serum.

Polyfluorenes with both alkyl and poly(ethylene glycol) (PEG) side-chains have been synthesised and processed into fluorescent nanoparticles via nanoprecipitation. The PEG/alkyl ratio is found to exert a significant influence over the size, polymer microstructure and optical properties of the resultant nanoparticles. This paves the way for a deeper understanding of how polymer structure can be manipulated to provide greater control over the nanoprecipitation process.

Polyfluorenes with pendant alkoxysilyl groups have been used to prepare inorganic-organic composite nanoparticles (diameter 80-220 nm) in which the conjugated polymer is dispersed within a silica matrix. Preparation of these nanoparticles is achieved by simultaneous nanoprecipitation of the conjugated polymer and hydrolysis/crosslinking of the alkoxysilyl groups under basic conditions. The composition of the nanocomposites is controlled by addition of an alkoxysilane monomer, tetramethylorthosilicate. The hybrid nanoparticles form highly stable dispersions in water and buffer (pH 9.2). The size of the nanoparticles can be tuned by varying the amount of the alkoxysilane monomer added during the nanoprecipitation process. Increasing the relative amount of alkoxysilane monomer also increases the proportion of polyfluorene chains that adopt the higher energy beta-phase conformation within the resultant nanoparticles. Nanoparticles with the highest silica content were found to have increased photoluminescence quantum yields. This work provides a controllable method for optimisation of the photophysical properties of light-emitting conjugated polymer nanoparticles via a simple aqueous processing technique.

The efficient transport of micron-sized beads into cells, via a non-endocytosis mediated mechanism, has only recently been described. As such there is considerable scope for optimization and exploitation of this procedure to enable imaging and sensing applications to be realized. Herein, we report the design, synthesis and characterization of fluorescent microsphere-based cellular delivery agents that can also carry biological cargoes. These core-shell polymer microspheres possess two distinct chemical environments; the core is hydrophobic and can be labeled with fluorescent dye, to permit visual tracking of the microsphere during and after cellular delivery, whilst the outer shell renders the external surfaces of the microspheres hydrophilic, thus facilitating both bioconjugation and cellular compatibility. Cross-linked core particles were prepared in a dispersion polymerization reaction employing styrene, divinylbenzene and a thiol-functionalized co-monomer. These core particles were then shelled in a seeded emulsion polymerization reaction, employing styrene, divinylbenzene and methacrylic acid, to generate orthogonally functionalized core-shell microspheres which were internally labeled via the core thiol moieties through reaction with a thiol reactive dye (DY630-maleimide). Following internal labeling, bioconjugation of green fluorescent protein (GFP) to their carboxyl-functionalized surfaces was successfully accomplished using standard coupling protocols. The resultant dual-labeled microspheres were visualized by both of the fully resolvable fluorescence emissions of their cores (DY630) and shells (GFP). In vitro cellular uptake of these microspheres by HeLa cells was demonstrated conventionally by fluorescence-based flow cytometry, whilst MTT assays demonstrated that 92% of HeLa cells remained viable after uptake. Due to their size and surface functionalities, these far-red-labeled microspheres are ideal candidates for in vitro, cellular delivery of proteins.

ADVERTISEMENT RETURN TO ISSUEPREVNoteNEXTStructural Analysis of Linear PEEK via MALDI-TOF Mass SpectrometryJonathan M. Behrendt†, Michael Benstead†‡, Adam Chaplin‡, Brian Wilson‡, and Michael L. Turner*†View Author Information† School of Chemistry, University of Manchester, Oxford Road, Manchester M13 9PL, U.K.‡ Victrex Technology Centre, Hillhouse International, Thorton-Cleveleys, Lancashire FY5 4QD, U.K.E-mail [email protected]; Tel 0161 275 4625.Cite this: Macromolecules 2011, 44, 22, 9054–9056Publication Date (Web):October 21, 2011Publication History Received11 July 2011Revised29 September 2011Published online21 October 2011Published inissue 22 November 2011https://pubs.acs.org/doi/10.1021/ma201580dhttps://doi.org/10.1021/ma201580dbrief-reportACS PublicationsCopyright © 2011 American Chemical SocietyRequest reuse permissionsArticle Views1177Altmetric-Citations5LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InRedditEmail Other access optionsGet e-AlertscloseSupporting Info (1)»Supporting Information Supporting Information SUBJECTS:Mass spectrometry,Oligomers,Polymer solutions,Polymers,Solvents Get e-Alerts

The naturally occurring reactive electrophilic species 12-oxo-phytodienoic acid (12-oxo-PDA) is a potent antifungal agent, whereas the plant growth regulator jasmonic acid, which is synthesized from 12-oxo-PDA, is ineffective. To address what structural features of the molecule endow it with antifungal activity, we synthesized a series of molecular mimics of 12-oxo-PDA varying in the length of the alkyl chain at its C-4 ring position. The octyl analogue (4-octyl cyclopentenone) was the most effective at suppressing spore germination and subsequent mycelial growth of a range of fungal pathogens and was particularly effective against Cladosporium herbarum and Botrytis cinerea, with minimum fungicidal concentrations in the range 100-200 µM. Introduction of a carboxyl group to the end of the chain, mimicking natural fatty acids, markedly reduced antifungal efficacy. Electrolyte leakage, indicative of membrane perturbation, was evident in both C. herbarum and B. cinerea exposed to 4-octyl cyclopentenone. Lipid composition analysis of the fungal spores revealed that those species with a high oil content, namely Fusarium oxysporum and Alternaria brassicicola, were less sensitive to 4-octyl cyclopentenone. The comparable hydrophobicity of 4-octyl cyclopentenone and 12-oxo-PDA accounts for the similar spore suppression activity of these two compounds. The relative ease of synthesis of 4-octyl cyclopentenone makes it an attractive compound for potential use as an antifungal agent.

Neural stem cells (NSC) are a valuable model system for understanding the intrinsic and extrinsic controls for self-renewal and differentiation choice. They also offer a platform for drug screening and neurotoxicity studies, and hold promise for cell replacement therapies for the treatment of neurodegenerative diseases. Fully exploiting the potential of this experimental tool often requires the manipulation of intrinsic cues of interest using transfection methods, to which NSC are relatively resistant. In this paper, we show that mouse and human NSC readily take up polystyrene-based microspheres which can be loaded with a range of chemical or biological cargoes. This uptake can take place in the undifferentiated stage without affecting NSC proliferation and their capacity to give rise to neurons and glia. We demonstrate that P-galactosidase-loaded microspheres could be efficiently introduced into NSC with no apparent toxic effect, thus providing proof-of-concept for the use of microspheres as an alternative biomolecule delivery system.