A single-step gold nanoparticle (AuNP)-based immunoassay is demonstrated in which the nanoparticle surface is tagged with short viral peptide epitopes. Antiviral antibodies with monoclonal specificity trigger nanoparticle aggregation yielding a colorimetric response that enables detection of antibodies in the low-nanomolar range within a few minutes. In silico insights into the interactions at the epitope−gold interface demonstrate that the conformational landscape exhibited by the epitopes is strongly influenced by the amino acid sequence and location of particular residues within the peptides. The conformation, orientation, and linker chemistry of the peptides affect the immune complex formation in nonintuitive ways that are, nevertheless, explained by a unique sterically kinetically driven aggregation mechanism. The rapid and specific performance of the AuNP immunoassay may have generic potential in point of care diagnostics and other laboratory routines.
This work describes a cell-based assay that does not depend on radioactivity or laboratory animals for the detection of ligands of angiotensin II type 1 receptor (AT1R). The assay makes use of stable transfected Chinese hamster ovary cells (CHO-AT1R) expressing the AT1R. A sequential saturation assay principle was used in which receptor binding sites of the CHO-AT1R cells are blocked by the analyte in a concentration-dependent manner. Afterwards, TAMRA-angiotensin II, a fluorescence-labeled ligand, was added to bind to the remaining free binding sites of the receptor. In consequence, the fluorescence signal determined is inversely proportional to the concentration of the analyte.
This work describes a method for surface regeneration of microfluidic microarray printheads through plasma techniques. Modification procedures were chosen in a way to obtain high reproducibility with a minimum of time consumption. The idea behind this is a complete regeneration of a microarray printhead before or after usage to achieve best printing results over a typical print job. A sequence of low-pressure oxygen-plasma and plasma polymerization with hexamethyldisiloxane (HMDSO) was used to regenerate printheads. Proof of the concept is given through quality control performed with a spotter implemented CCD camera, contact angle measurements and a typical hybridization experiment. Stable printing results were obtained over 3000 activations showing that the presented method is suitable for treatment of microarray printheads.
During the last decade microarrays have become a powerful analytical tool. Commonly microarrays are produced in a non-contact manner using silicone printheads. However, silicone printheads are expensive and not able to be used as a disposable. Here, we show the development and functional characterization of 8-channel plastic microarray printheads that overcome both disadvantages of their conventional silicone counterparts. A combination of injection-molding and laser processing allows us to produce a high quantity of cheap, customizable and disposable microarray printheads. The use of plastics (e.g., polystyrene) minimizes the need for surface modifications required previously for proper printing results. Time-consuming regeneration processes, cleaning procedures and contaminations caused by residual samples are avoided. The utilization of plastic printheads for viscous liquids, such as cell suspensions or whole blood, is possible. Furthermore, functional parts within the plastic printhead (e.g., particle filters) can be included. Our printhead is compatible with commercially available TopSpot devices but provides additional economic and technical benefits as compared to conventional TopSpot printheads, while fulfilling all requirements demanded on the latter. All in all, this work describes how the field of traditional microarray spotting can be extended significantly by low cost plastic printheads.
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