Reusable surface amplified nanobiosensor for the sub PFU/mL level detection of airborne virus.

We developed a reusable surface-amplified nanobiosensor for monitoring airborne viruses with a sub-PFU/mL level detection limit. Here, sandwich structures consisted of magnetic particles functionalized with antibodies, target viruses, and alkaline phosphatases (ALPs) were formed, and they were magnetically concentrated on Ni patterns near an electrochemical sensor transducer. Then, the electrical signals from electrochemical markers generated by ALPs were measured with the sensor transducer, enabling highly-sensitive virus detection. The sandwich structures in the used sensor chip could be removed by applying an external magnetic field, and we could reuse the sensor transducer chip. As a proof of concepts, the repeated detection of airborne influenza virus using a single sensor chip was demonstrated with a detection limit down to a sub-PFU/mL level. Using a single reusable sensor transducer chip, the hemagglutinin (HA) of influenza A (H1N1) virus with different concentrations were measured down to 10 aM level. Importantly, our sensor chip exhibited reliable sensing signals even after more than 18 times of the repeated HA sensing measurements. Furthermore, airborne influenza viruses collected from the air could be measured down to 0.01 PFU/mL level. Interestingly, the detailed quantitative analysis of the measurement results revealed the degradation of HA proteins on the viruses after the air exposure. Considering the ultrasensitivity and reusability of our sensors, it can provide a powerful tool to help preventing epidemics by airborne pathogens in the future.

Calibration curve-free electrochemical quantitation by micro-nano multi-scale gap devices.

Quantitation without relying on the calibration curve has long been an issue of overcoming analytical problems accompanied with the inherent limitations of the calibration curve fitting errors. Here, we report on a calibration curve-free method for electrochemical quantitation based on a multi-scale gap device (MGD). The MGD is an integrated device having a series of interdigitated electrodes (IDE) with micro-to-nano gap distances. The device shows a gap-dependent redox current of the analyte when subjected to the electrochemical cycling between the two facing electrodes of its componential IDEs. Based on the fact that the current increases as the gap distance decreases, the analyte concentration could be directly estimated: the rate of increase in the current was directly proportional to the analyte concentration. The calibration curve was not necessary for the quantitation. The accuracy of this MGD approach was better than that of an IDE collection of the same gap distance, which was deteriorated at the larger gap distances particularly. The MGD-based quantitation of dopamine, potassium ferricyanide, and aminophenol was demonstrated in a relatively broad range of concentrations (100 nM-5 mM).

Electrochemical microgap immunosensors for selective detection of pathogenic Aspergillus niger.

Aspergillus niger (A. niger) is a well-known allergenic, harmful fungus in the indoor environment that can cause asthmatic symptoms and atopy. Previous immunosensing approach suffers from an insufficient detection limit, mainly because there are no techniques for target amplification. We report an electrochemical immunosensor that selectively quantifies the A. niger based on the detection of extracellular proteins by using a specific interaction with antibody. The sensor was designed to show a decrease in redox current upon binding of the antigens secreted from A. niger onto an antibody-immobilized surface between the interdigitated electrodes. The extracellular proteins were profiled by LC-MS/MS to identify the antigens existing in the A. niger solution. Since the targets of the sensor are the proteins, its sensitivity and selectivity remain almost intact even after filtration of the spores. It was also found that the use of secretion promoter in the sampling stage greatly improved the sensor's limit of detection (LOD) for the spores. By this, the LOD was lowered by a few orders of magnitude so as to reach the value as low as ~101 spores/mL.

Potential Role of Soluble Metal Impurities in the Acute Lung Inflammogenicity of Multi-Walled Carbon Nanotubes.

Multi-walled carbon nanotubes (MWCNTs) have variable metal impurities, but little is known about the impact of soluble metal impurities on the toxicity of MWCNTs. Here, we evaluated the role of soluble metal impurities to the acute inflammogenic potential of MWCNTs, using five types of high purity MWCNTs (>95%). MWCNTs and their soluble fractions collected at 24 h after incubation in phosphate-buffered saline showed diverse metal impurities with variable concentrations. The fiber-free soluble fractions produced variable levels of reactive oxygen species (ROS), and the iron level was the key determinant for ROS production. The acute inflammation at 24 h after intratracheal instillation of MWCNTs to rats at 0.19, 0.63, and 1.91 mg MWCNT/kg body weight (bw) or fiber-free supernatants from MWCNT suspensions at 1.91 and 7.64 mg MWCNT/kg bw showed that the number of granulocytes, a marker for acute inflammation, was significantly increased with a good dose-dependency. The correlation study showed that neither the levels of iron nor the ROS generation potential of the soluble fractions showed any correlations with the inflammogenic potential. However, the total concentration of transition metals in the soluble fractions showed a good correlation with the acute lung inflammogenic potential. These results implied that metal impurities, especially transitional metals, can contribute to the acute inflammogenic potential of MWCNTs, although the major parameter for the toxicity of MWCNTs is size and shape.

Threshold Rigidity Values for the Asbestos-like Pathogenicity of High-Aspect-Ratio Carbon Nanotubes in a Mouse Pleural Inflammation Model.

The qualitative and quantitative evaluation of the physicochemical parameters associated with the pathogenicity of high-aspect-ratio nanomaterials is important for comprehensive regulation efforts and safety-by-design approaches. Here, we report quantitative data on the correlations between the rigidity of these nanomaterials and toxicity endpoints in vitro and in vivo. As measured by new ISO standards published in 2017, rigidity shows a strong positive correlation with inflammogenic potential, as indicated by inflammatory cell counts and IL-1ß (a biomarker for frustrated phagocytosis) levels in both the acute and chronic phases. In vitro experiments using differentiated THP-1 cells find that only highly rigid multiwalled carbon nanotubes (MWCNTs) and asbestos fibers lead to piercing and frustrated phagocytosis. Thus, this study suggests a bending ratio of 0.97 and a static bending persistence length of 1.08 as threshold rigidity values for asbestos-like pathogenicity. However, additional research using MWCNTs with rigidity values that lie between those of non-inflammogenic ( Db = 0.66 and SBPL = 0.87) and inflammogenic fibers ( Db = 0.97 and SBPL = 1.09) is required to identify more accurate threshold values, which would be useful for comprehensive regulation and safety-by-design approaches based on MWCNTs.

Long dsRNA-mediated RNA interference and immunostimulation: a targeted delivery approach using polyethyleneimine based nano-carriers.

RNA oligonucleotides capable of inducing controlled immunostimulation combined with specific oncogene silencing via an RNA interference (RNAi) mechanism provide synergistic inhibition of cancer cell growth. With this concept, we previously designed a potent immunostimulatory long double stranded RNA, referred to as liRNA, capable of executing RNAi mediated specific target gene silencing. In this study, we developed a highly effective liRNA based targeted delivery system to apply in the treatment of glioblastoma multiforme. A stable nanocomplex was fabricated by complexing multimerized liRNA structures with cross-linked branched poly(ethylene imine) (bPEI) via electrostatic interactions. We show clear evidence that the cross-linked bPEI was quite effective in enhancing the cellular uptake of liRNA on U87MG cells. Moreover, the liRNA-PEI nanocomplex provided strong RNAi mediated target gene silencing compared to that of the conventional siRNA-PEI complex. Further, the bPEI modification strategy with specific ligand attachment assisted the uptake of the liRNA-PEI complex on the mouse brain endothelial cell line (b.End3). Such delivery systems combining the beneficial elements of targeted delivery, controlled immunostimulation, and RNAi mediated target silencing have immense potential in anticancer therapy.

Calcination-temperature-Dependent shape and crystal structure of TiO2 nanomaterials synthesized by hard-template method.

The shape and crystal structure of TiO2 nanomaterials synthesized by the hard-template method can be controlled by simply changing the calcination temperature. In this work, SiO2 nanoparticles were used as a hard template and TiO2 was coated onto the surface of the silica core, resulting in core-shell nanoparticles, which were then calcined at various temperatures to induce shape transformation and crystallization of the TiO2 shell. After etching of the silica cores, spherical hollow nanocapsules with anatase crystal phase were obtained by calcination at 400-1000 degrees C, while urchin-like hollow capsules and small-sized particulates were obtained at temperatures below 400 degrees C and above 1000 degrees C, respectively. The core-shell nanoparticles exhibited greatly enhanced anatase phase stability (up to approximately 1200 degrees C), which was attributable to the effect of the core material. The phase stability was found to be dependent on the shell thickness of the nanocapsules, also supporting the effect of the core material.