Diagnosis of Toxoplasmosis Using a Synthetic Glycosylphosphatidylinositol Glycan

Around 2 billion people worldwide are infected with the apicomplexan parasite Toxoplasma gondii which induces a variety of medical conditions. For example, primary infection during pregnancy can result in fetal death or mental retardation of the child. Diagnosis of acute infections in pregnant women is challenging but crucially important as the drugs used to treat T. gondii infections are potentially harmful to the unborn child. Better, faster, more reliable, and cheaper means of diagnosis by using defined antigens for accurate serological tests are highly desirable. Synthetic pathogen‐specific glycosylphosphatidylinositol (GPI) glycan antigens are diagnostic markers and have been used to distinguish between toxoplasmosis disease states using human sera.     

Rupture force of single supramolecular bonds in associative polymers by AFM at fixed loading rates

Atomic-force-microscopy-based single-molecule force spectroscopy (AFM-SMFS) was used to study the bond strength of self-complementary hydrogen-bonded complexes based on the 2-ureido-4[1H]-pyrimidinone (UPy) quadruple H-bond motif in hexadecane (HD). The unbinding force corresponding to single UPy-UPy dimers was investigated at a fixed piezo retraction rate in the nonequilibrium loading rate regime. The rupture force of bridging supramolecular polymer chains formed between UPy-functionalized substrates and AFM tips in the presence of a bis-UPy derivative was found to decrease with increasing rupture length. The rupture length was identified as the chain length of single, associating polymers, which allowed us to determine the number of supramolecular bonds (N) at rupture. The rupture force observed as a function of N was in quantitative agreement with the theory on uncooperative bond rupture for supramolecular linkages switched in a series. Hence, the value of the dimer equilibrium constant Keq=(1.3+/-0.5) x 10(9) M(-1), which is in good agreement with previously estimated values, was obtained by SMFS of supramolecular polymers at a single loading rate.     

Translation of pseudo-cross-conjugation into chemistry: cycloadditions of mesomeric betaines to the new ring system spiro[indazole-3,3'-pyrrole

Indazolium-3-amidates (X-ray analysis), readily available on trapping the N-heterocyclic carbene indazol-3-ylidene with isocyanates, underwent [3+2]-cycloadditions with activated triple bonds to spiro[indazole-3,3'-pyrroles]. A combination of NMR techniques such as heteronuclear single quantum coherence (HSQC), heteronuclear multiple bond correlation (HMBC), nuclear Overhauser enhancement spectroscopy (NOESY), and 1H/15N correlations were applied to elucidate the structures of the cycloadducts.     

Influence of size and functionality of polymeric nanoparticles on the adsorption behavior of sodium dodecyl sulfate as detected by isothermal titration calorimetry

Isothermal titration calorimetry was used to monitor the adsorption of the surfactant sodium dodecylsulfate (SDS) on different sized pure and carboxy functionalized polystyrene nanoparticles prepared by the mini-emulsion process. The ITC experiment gives, additionally to the CMC values, information about the interaction of the surfactant molecules to the particle’s surface due to the particle surface properties. The adsorption heat depends on the chemical composition of the polymer as well on the particle size. It also provides information about the surface coverage with surfactant and the number of additional adsorbed molecules per particle until full coverage by surfactant is obtained. The surfactant adsorption increases from 0.3 molecules per nm² for 50 nm to 8.5 molecules per nm² for carboxy functionalized particles with diameters larger than 160 nm. The area A _Surf-dens after the adsorption process gives information about the packing density of surfactant molecules on the particles in dependence of carboxy groups: an increasing number of carboxylic groups decreases the area occupied per SDS molecule. The adsorption process was also monitored by zeta potential measurements, where an increasing potential during the adsorption was detected.     

Synthesis and characterization of dually labeled Pickering-type stabilized polymer nanoparticles in a downscaled miniemulsion system

Dual fluorescently labeled polymer particles were prepared in a downscaled Pickering-type miniemulsion system. Stable dispersions were obtained and the size of the hybrid particles could be varied between ca. 180 and 430 nm. Silica nanoparticles were employed as sole emulsifier, which were labeled by a fluorescein dye (FITC) or (encapsulated) quantum dots, and the polymer core was labeled by a perylene derivative. Downscaling of the Pickering-type miniemulsion system is intriguing by itself as it allows the use of precious nanoparticles as emulsifiers. Here, silica particles with a fluorescent core and an overall diameter between 20 and 40 nm were prepared and employed as stabilizer. The dual excitation and emission of both dyes was tested by fluorescence measurements and confocal laser scanning microscopy (cLSM).     

Quantum dots as simultaneous acceptors and donors in time-gated Förster resonance energy transfer relays: characterization and biosensing

The unique photophysical properties of semiconductor quantum dot (QD) bioconjugates offer many advantages for active sensing, imaging, and optical diagnostics. In particular, QDs have been widely adopted as either donors or acceptors in F?rster resonance energy transfer (FRET)-based assays and biosensors. Here, we expand their utility by demonstrating that QDs can function in a simultaneous role as acceptors and donors within time-gated FRET relays. To achieve this configuration, the QD was used as a central nanoplatform and coassembled with peptides or oligonucleotides that were labeled with either a long lifetime luminescent terbium(III) complex (Tb) or a fluorescent dye, Alexa Fluor 647 (A647). Within the FRET relay, the QD served as a critical intermediary where (1) an excited-state Tb donor transferred energy to the ground-state QD following a suitable microsecond delay and (2) the QD subsequently transferred that energy to an A647 acceptor. A detailed photophysical analysis was undertaken for each step of the FRET relay. The assembly of increasing ratios of Tb/QD was found to linearly increase the magnitude of the FRET-sensitized time-gated QD photoluminescence intensity. Importantly, the Tb was found to sensitize the subsequent QD-A647 donor-acceptor FRET pair without significantly affecting the intrinsic energy transfer efficiency within the second step in the relay. The utility of incorporating QDs into this type of time-gated energy transfer configuration was demonstrated in prototypical bioassays for monitoring protease activity and nucleic acid hybridization; the latter included a dual target format where each orthogonal FRET step transduced a separate binding event. Potential benefits of this time-gated FRET approach include: eliminating background fluorescence, accessing two approximately independent FRET mechanisms in a single QD-bioconjugate, and multiplexed biosensing based on spectrotemporal resolution of QD-FRET without requiring multiple colors of QD.