13 research outputs found
ThiolâClick Based Polyglycerol Hydrogels as Biosensing Platform with In Situ Encapsulated Streptavidin Probes
An in situ streptavidinâencapsulated hydrogel based on dendritic polyglycerol (dPG) which is functionalized with either an acrylate, allyl or acrylamide group and dithiolated polyethylene glycol (PEG) is constructed via a thiolâclick chemistry approach and is investigated for biosensing applications. The hydrogel platform is screened for the encapsulation and release efficiencies of the model protein streptavidin under varying physicochemical conditions, for example, crosslinking chemistry reactions, the molar ratio between the two gel components, macromonomer concentrations or pHâvalues. By that, tailorâmade hydrogels can be developed, which are able to encapsulate or release the model protein for several days based on its modality. Furthermore, the accessible binding site of encapsulated streptavidin or in other words, the biotinâbinding performance is quantified, and the stability of the various hydrogel types is studied by rheology measurements, 1H NMR, gel permeation chromatography (GPC), and mass loss experiments
Precise supramolecular control of surface coverage densities on polymer micro- and nanoparticles
We report herein the controlled surface functionalization of micro- and nanoparticles by supramolecular
hostâguest interactions. Our idea is to exploit the competition of two high-affinity guests for binding to
the surface-bound supramolecular host cucurbit[7]uril (CB7). To establish our strategy, surface azide
groups were introduced to hard-sphere (poly)methylmethacrylate particles with a grafted layer of
poly(acrylic acid), and a propargyl derivative of CB7 was coupled to the surface by click chemistry. The
amount of surface-bound CB7 was quantified with the high-affinity guest aminomethyladamantane
(AMADA), which revealed CB7 surface coverage densities around 0.3 nmol cm 2 indicative of a 3D layer
of CB7 binding sites on the surface. The potential for surface functionalization was demonstrated with an
aminoadamantane-labeled rhodamine (Ada-Rho) as a second high-affinity guest. Simultaneous
incubation of CB7-functionalized particles with both high-affinity guests, AMADA and Ada-Rho, revealed
a simple linear relationship between the resulting surface coverage densities of the model fluorescent
dye and the mole fraction of Ada-Rho in the incubation mixture. This suggests a highly modular
supramolecular strategy for the stable immobilization of application-relevant molecules on particle
surfaces and a precise control of their surface coverage densities.The financial support from the Deutsche Forschungsgemeinschaft (HE 5967/4-1 and NA 686/11-1), the China Scholarship Council (S. Z.), the Ministerio de Economia, Industria y Competitividad, Madrid (grant CTQ2014-54729-C2-1-P and predoctoral contract BES-2015-074458 for Z. D.), and the Ministerio de Ciencia, Innovacion y Universidades, Madrid (grant CTQ2017-89832-P) is gratefully acknowledged
Bioorthogonal in Situ Hydrogels Based on Polyether Polyols for New Biosensor Materials with High Sensitivity
Both
noncovalent and covalent encapsulations of active biomolecules,
for example, proteins and oligonucleotides, for a new biosensor matrix
in an in situ bioorthogonal hydrogel formation via a strain-promoted
azideâalkyne cycloaddition reaction were investigated. Unspecific
interaction between the gel and the biomolecules as well as protein
denaturation was prevented by the bioorthogonal gel components, which
ensure a uniform aqueous environment in the hydrogel network. No leaching
of the active biomolecules was observed. Additionally, a much higher
and also adjustable loading of biomolecules in the hydrogel matrix
was achieved compared to conventional biosensor surfaces, where the
sensor molecules are immobilized on monolayers (2D surfaces) or brushlike
structures (3D surfaces). Spotting experiments of the hydrogel confirm
the possibility to use this new surface for microarray-based multiplex
applications which require very high signal-to-noise ratios
Close Spectroscopic Look at Dye-Stained Polymer Microbeads
Dye-stained
micrometer-sized polymer beads are important tools
in the life sciences with applications in biomedical, biochemical,
and clinical research. Here, bead-based assays are increasingly used,
for example, in DNA sequencing and the detection of autoimmune diseases
or pathogenic microorganisms. Moreover, stained beads are employed
as calibration tools for fluorescence microscopy and flow cytometry
methods with increasing complexity. To address the requirements concerning
the relevant fluorescence features, the spectroscopic properties of
representative polymer beads with diameters ranging from about 1 to
10 Îźm stained with varying concentrations of rhodamine 6G were
systematically assessed. The observed dependence of the spectral properties,
fluorescence decay kinetics, and fluorescence quantum yields on bead
size and dye loading concentration is attributed to different fluorescence
characteristics of fluorophores located in the particle core and near-surface
dye molecules. Supported by the fluorescence anisotropy measurements,
the origin of the observed alteration of fluorescence features is
ascribed to a combination of excitation energy transfer and polarity-related
effects that are especially pronounced at the interface of the bead
and the surrounding medium. The results of our studies underline the
need to carefully control and optimize all parameters that can affect
the fluorescence properties of the dye-stained beads
Scope and Limitations of Surface Functional Group Quantification Methods: Exploratory Study with Poly(acrylic acid)-Grafted Micro- and Nanoparticles
The amount of grafted polyÂ(acrylic acid) on polyÂ(methyl
methacrylate)
micro- and nanoparticles was quantified by conductometry, <sup>13</sup>C solid-state NMR, fluorophore labeling, a supramolecular assay based
on high-affinity binding of cucurbit[7]Âuril, and two colorimetric
assays based on toluidine blue and nickel complexation by pyrocatechol
violet. The methods were thoroughly validated and compared with respect
to reproducibility, sensitivity, and ease of use. The results demonstrate
that only a small but constant fraction of the surface functional
groups is accessible to covalent surface derivatization independently
of the total number of surface functional groups, and different contributing
factors are discussed that determine the number of probe molecules
which can be bound to the polymer surface. The fluorophore labeling
approach was modified to exclude artifacts due to fluorescence quenching,
but absolute quantum yield measurements still indicate a major uncertainty
in routine fluorescence-based surface group quantifications, which
is directly relevant for biochemical assays and medical diagnostics.
Comparison with results from protein labeling with streptavidin suggests
a porous network of polyÂ(acrylic acid) chains on the particle surface,
which allows diffusion of small molecules (cutoff between 1.6 and
6.5 nm) into the network
Politicizing Europe: the challenge of executive discretion
Political decision-making in the Euro-crisis has relied heavily on executive discretion, exercised at speed and rationalised with reference to the pressing demands of emergency. This paper explores the challenges raised for political opposition, notably challenges of a temporal kind. With its deviations from policy and procedural norms, discretionary politics tends towards a politics without rhythm, leading to major asymmetries between decisionmakers and voices of opposition. These centre on issues of timing and the ability to identify authorship and content of decisions. Such asymmetries arguably correspond to an underlying one between the temporality of political decision-making and of contemporary finance capitalism, with agents of the former increasingly inclined to pursue âfast policyâ as a means to keep pace. A democratic response is likely to involve strengthening and synchronising the rhythms of parliamentary politics, as well as being receptive to forms of opposition less reliant on the rhythms that discretion subverts