Quantitative Detection of Digoxin in Plasma Using Small-Molecule Immunoassay in a Recyclable Gravity-Driven Microfluidic Chip.

Immunoassays are critical for clinical diagnostics and biomedical research. However, two major challenges remaining in conventional immunoassays are precise quantification and development of immunoassays for small-molecule detection. Here, a two signal-mode small-molecule immunoassay containing an internal reference that provides high stability and reproducibility compared to conventional small-molecule immunoassays is presented.

Imidazole carbamate probes for affinity guided azide-transfer to metal-binding proteins.

Probes for affinity guided conjugation have shown great promise for the preparation of high-quality protein conjugates. However, such probes are often limited in their protein scope. Here, we demonstrate the synthesis and use of imidazole carbamate probes for affinity guided conjugation of an azide handle to a wide variety of metal binding proteins such as antibodies, enzymes, nanobodies and His6-tagged proteins.

Dimethyl fumarate is an allosteric covalent inhibitor of the p90 ribosomal S6 kinases.

Dimethyl fumarate (DMF) has been applied for decades in the treatment of psoriasis and now also multiple sclerosis. However, the mechanism of action has remained obscure and involves high dose over long time of this small, reactive compound implicating many potential targets. Based on a 1.

Active Intracellular Delivery of a Cas9/sgRNA Complex Using Ultrasound-Propelled Nanomotors.

Direct and rapid intracellular delivery of a functional Cas9/sgRNA complex using ultrasound-powered nanomotors is reported. The Cas9/sgRNA complex is loaded onto the nanomotor surface through a reversible disulfide linkage. A 5 min ultrasound treatment enables the Cas9/sgRNA-loaded nanomotors to directly penetrate through the plasma membrane of GFP-expressing B16F10 cells.

AFM Imaging of Hybridization Chain Reaction Mediated Signal Transmission between Two DNA Origami Structures.

Signal transfer is central to the controlled exchange of information in biology and advanced technologies. Therefore, the development of reliable, long-range signal transfer systems for artificial nanoscale assemblies is of great scientific interest. We have designed such a system for the signal transfer between two connected DNA nanostructures, using the hybridization chain reaction (HCR).

Phosphines as Efficient Dioxygen Scavengers in Nitrous Oxide Sensors.

A current challenge for development of amperometric sensors for the greenhouse gas nitrous oxide (NO) is their sensitivity toward dioxygen and trace water. The need for aqueous dioxygen scavengers in front of the sensor implies a background signal from penetrating water vapor. In this paper, we introduce substituted phosphines as dioxygen scavengers and demonstrate the application in a dioxygen-insensitive NO sensor.

Dynamic Chemistry of Disulfide Terminated Oligonucleotides in Duplexes and Double-Crossover Tiles.

Designed nanostructures formed by self-assembly of multiple DNA strands suffer from low stability at elevated temperature and under other denaturing conditions. Here, we propose a method for covalent coupling of DNA strands in such structures by the formation of disulfide bonds; this allows disassembly of the structure under reducing conditions. The dynamic chemistry of disulfides and thiols was applied to crosslink DNA strands with terminal disulfide modifications.

Programmed Switching of Single Polymer Conformation on DNA Origami.

DNA nanotechnology offers precise geometrical control of the positioning of materials, and it is increasingly also being used in the development of nanomechanical devices. Here we describe the development of a nanomechanical device that allows switching of the position of a single-molecule conjugated polymer. The polymer is functionalized with short single-stranded (ss) DNA strands that extend from the backbone of the polymer and serve as handles.

Synthesis, dynamic combinatorial chemistry, and PCR amplification of 3'-5' and 3'-6' disulfide-linked oligonucleotides.

Disulfide dithymidines linked 3'-5' or 3'-6' were synthesized and incorporated into oligonucleotides through a combined phosphotriester and phosphoramidite solid-phase oligonucleotide synthesis approach. The disulfide links are cleaved and formed reversibly in the presence of thiols and oligonucleotides. This link was shown to be sequence-adaptive in response to given templates in the presence of mercaptoethanol.

Isothermal hybridization kinetics of DNA assembly of two-dimensional DNA origami.

The Watson-Crick base-pairing with specificity and predictability makes DNA molecules suitable for building versatile nanoscale structures and devices, and the DNA origami method enables researchers to incorporate more complexities into DNA-based devices. Thermally controlled atomic force microscopy in combination with nanomechanical spectroscopy with forces controlled in the pico Newton (pN) range as a novel technique is introduced to directly investigate the kinetics of multistrand DNA hybridization events on DNA origami nanopores under defined isothermal conditions. For the synthesis of DNA nanostructures under isothermal conditions at 60 °C, a higher hybridization rate, fewer defects, and a higher stability are achieved compared to room-temperature studies.

Self-assembled nanoparticles of modified-chitosan conjugates for the sustained release of DL-α-tocopherol.

Synthetic O6-succinylated chitosan and commercial glycol chitosan were covalently linked to dl-α-tocopheryl monoesters for controlled release of vitamin E. These conjugates formed self-assembled nanoparticles in aqueous solution with 254-496 nm mean diameters and dl-α-tocopherol contents between 27 and 39% (w/w). The particles appeared as 40-75 nm almost spherical nanoparticles when studied by scanning and transmission electron microscopy upon drying.

N,O6-partially acetylated chitosan nanoparticles hydrophobically-modified for controlled release of steroids and vitamin E.

Diosgenin, two synthetic analogs of brassinosteroids, testosterone and dl-α-tocopherol were covalently linked to synthetic water-soluble N,O6-partially acetylated chitosan, for their controlled release. Drug linking was confirmed by FTIR spectroscopy and proton NMR. Conjugates were also characterized by differential scanning calorimetry and wide-angle X-ray diffraction.

Direct visualization of transient thermal response of a DNA origami.

The DNA origami approach enables the construction of complex objects from DNA strands. A fundamental understanding of the kinetics and thermodynamics of DNA origami assembly is extremely important for building large DNA structures with multifunctionality. Here both experimental and theoretical studies of DNA origami melting were carried out in order to reveal the reversible association/disassociation process.

Synthesis of dopamine and serotonin derivatives for immobilization on a solid support.

The two important neurotransmitters dopamine and serotonin are synthesized with short PEG tethers and immobilized on a magnetic solid support. The tether is attached to the aromatic moiety of the neurotransmitters to conserve their original functional groups. This approach causes minimal alteration of the original structure with the aim of optimizing the immobilized neurotransmitters for aptamer selection by SELEX.

Single molecule microscopy methods for the study of DNA origami structures.

Single molecule microscopy techniques play an important role in the investigation of advanced DNA structures such as those created by the DNA origami method. Three single molecule microscopy techniques are particularly interesting for the investigation of complex self-assembled three-dimensional (3D) DNA nanostructures, namely single molecule fluorescence microscopy, atomic force microscopy (AFM), and cryogenic transmission electron microscopy (cryo-EM). Here we discuss the strengths of these three techniques and demonstrate how their interplay can yield very important and unique new insights into the structure and conformation of advanced biological nanostructures.

Functional patterning of DNA origami by parallel enzymatic modification.

We demonstrate here a rapid and cost-effective technique for nanoscale patterning of functional molecules on the surface of a DNA origami. The pattern is created enzymatically by transferring a functionalized dideoxynucleotide to the 3'-end of an arbitrary selected set of synthetic DNA oligonucleotides positioned approximately 6 nm apart in a 70 × 100 nm(2) rectangular DNA origami. The modifications, which are performed in a single-tube reaction, provide an origami surface modified with a variety of functional groups including chemical handles, fluorescent dyes, or ligands for subsequent binding of proteins.

Site-specific chemical labeling of long RNA molecules.

Site-specific labeling of RNA molecules is a valuable tool for studying their structure and function. Here, we describe a new site-specific RNA labeling method, which utilizes a DNA-templated chemical reaction to attach a label at a specific internal nucleotide in an RNA molecule. The method is nonenzymatic and based on the formation of a four-way junction, where a donor strand is chemically coupled to an acceptor strand at a specific position via an activated chemical group.

Single-molecule chemical reactions on DNA origami.

DNA nanotechnology and particularly DNA origami, in which long, single-stranded DNA molecules are folded into predetermined shapes, can be used to form complex self-assembled nanostructures. Although DNA itself has limited chemical, optical or electronic functionality, DNA nanostructures can serve as templates for building materials with new functional properties. Relatively large nanocomponents such as nanoparticles and biomolecules can also be integrated into DNA nanostructures and imaged.

A yoctoliter-scale DNA reactor for small-molecule evolution.

The center of DNA three-way junctions, constituting a yoctoliter (10(-24) L) volume, is applied as an efficient reactor to create DNA-encoded libraries of chemical products. Amino acids and short peptides are linked to oligonucleotides via cleavable and noncleavable linkers. The oligonucleotide sequences contain two universal assembling domains at the center and a distal codon sequence specific for the attached building block.

Preparation temperature dependence of size and polydispersity of alkylthiol monolayer protected gold clusters.

The influence of preparation temperature on the size and size distribution of dodecylthiol monolayer protected gold clusters was studied. The monolayer protected clusters (MPCs) were synthesized by two different variations of the Brust-Schiffrin procedure. In all of the experiments, the stoichiometry of the reactants dodecylthiol, HAuCl(4), and sodium borohydride was kept constant, while the temperature was varied in the range of -18 to +90 degrees C.