Cost-effective production of tag-less recombinant protein in Nicotiana benthamiana.

Plants have recently received a great deal of attention as a means of producing recombinant proteins. Despite this, a limited number of recombinant proteins are currently on the market and, if plants are to be more widely used, a cost-effective and efficient purification method is urgently needed. Although affinity tags are convenient tools for protein purification, the presence of a tag on the recombinant protein is undesirable for many applications.

Nanoscale analysis of a functionalized polythiophene surface by adhesion mapping.

Functionalized ethylenedioxythiophene (EDOT) monomers, hydroxymethyl EDOT (EDOT-OH), and zwitterionic phosphorylcholine EDOT (EDOT-PC) were electropolymerized to prepare the homopolymers poly(EDOT-OH) and poly(EDOT-PC), and mixtures of these monomers were used to produce the copolymer poly(EDOT-OH)-co-poly(EDOT-PC). Force-extension-curve-based atomic force microscopy (AFM) was utilized to analyze the surfaces of the films. The PEDOT-OH film yielded force-extension curves for short stretching, and the PEDOT-PC film yielded curves for long stretching.

Phosphokinase antibody arrays on dendron-coated surface.

Monitoring protein phosphorylation at the cellular level is important to understand the intracellular signaling. Among the phosphoproteomics methods, phosphokinase antibody arrays have emerged as preferred tools to measure well-characterized phosphorylation in the intracellular signaling. Here, we present a dendron-coated phosphokinase antibody array (DPA) in which the antibodies are immobilized on a dendron-coated glass slide.

Direct quantitative analysis of HCV RNA by atomic force microscopy without labeling or amplification.

Force-based atomic force microscopy (AFM) was used to detect HCV (hepatitis C virus) RNA directly and to quantitatively analyse it without the need for reverse transcription or amplification. Capture and detection DNA probes were designed. The former was spotted onto a substrate with a conventional microarrayer, and the latter was immobilized on an AFM probe.

"Seeing and counting" individual antigens captured on a microarrayed spot with force-based atomic force microscopy.

The mapping capability of atomic force microscopy (AFM) enabled us to see captured prostate-specific antigens (PSAs) on a spot microarrayed with the corresponding antibody and count the number of the antigens in a submicrometer area. To enhance the reliability and the reproducibility of the approach, a third-generation dendron was employed for the surface treatment. The specific force between the captured PSA and the detection antibody (5A6) was measured after cross-linking, and the mean unbinding force was 56 +/- 2 pN.

Nanotechnology for early cancer detection.

Vast numbers of studies and developments in the nanotechnology area have been conducted and many nanomaterials have been utilized to detect cancers at early stages. Nanomaterials have unique physical, optical and electrical properties that have proven to be very useful in sensing. Quantum dots, gold nanoparticles, magnetic nanoparticles, carbon nanotubes, gold nanowires and many other materials have been developed over the years, alongside the discovery of a wide range of biomarkers to lower the detection limit of cancer biomarkers.

Dendron-modified polystyrene microtiter plate: surface characterization with picoforce AFM and influence of spacing between immobilized amyloid beta proteins.

A polystyrene microtiter plate was coated with a molecular layer of a cone-shaped dendron as a means of providing proper spacing between immobilized biomolecules. For the coating preparation, di(ethylene glycol) vinyl ether was grafted onto the surface of the microtiter plate by a plasma process followed by self-assembly of a second-generation dendron (9-acid) or a third-generation dendron (27-acid). Contact angle analysis revealed a pronounced increase in the hydrophilicity upon plasma grafting, while the hydrophilicity reverted/decreased after dendron immobilization.

Potential lead for an Alzheimer drug: a peptide that blocks intermolecular interaction and amyloid beta protein-induced cytotoxicity.

A peptide chAbeta30-16 (15-mer; CTFVRTHIFCKEHQF) was designed to bind to a region encompassing the entire polymerization-related (16KLVFF20) and part of the polymerization and toxicity-related (25GSNKGAIIGLM35) regions of amyloid beta-protein, Abeta1-42 by a hydropathic complementary approach. This peptide efficiently binds to Abeta and blocks intermolecular interaction and the formation of Abeta aggregates. In addition, the peptide neutralizes the cell toxicity of Abeta fibrils.

Expression and functional reconstitution of a recombinant antibody (Fab') specific for human apolipoprotein B-100.

We have cloned and constructed plasmid vectors, pETB23H and pETB23L, for bacterial expression of heavy (H) and light (L) chain cDNAs of Fab' of mAbB23 a monoclonal antibody specific to human plasma apolipoprotein (apo) B-100. The H- and L-chains were expressed as insoluble inclusion bodies in the cytoplasm of Escherichia coli. The inclusion bodies of both chains were isolated from the cell lysate, solubilized in 6 M guanidium-HCl, and mixed in equal molar amounts.

Bacterial expression and in vitro refolding of a single-chain fv antibody specific for human plasma apolipoprotein B-100.

From the cloned heavy and light chains of a murine monoclonal antibody (mAbB23) which is specific for human apolipoprotein (apo) B-100 of plasma low-density lipoproteins, a vector was designed for expression of a single-chain antibody (scFv) of mAbB23 in Escherichia coli. The expression vector was constructed so that the scFv gene (V(L)-linker-V(H)) was expressed under the control of the T7 promoter. The inclusion body of scFv was isolated from E.