Evaluation and selection of potent fluorescent immunosensors by combining fluorescent peptide and nanobodies displayed on yeast surface

Quenchbody (Q-body) is a quench-based fluorescent immunosensor labeled with fluorescent dye(s) near the antigen-binding site of an antibody. Q-bodies can detect a range of target molecules rapidly and directly. However, because Q-bodies show different antigen responses depending on the antibody used, time-consuming optimization of the Q-body structure is often necessary, and a high-throughput screening method for discriminating and selecting good Q-bodies is required. Here, we aimed to develop a molecular display method of nanobody-based “mini Q-bodies” by combining yeast surface display and coiled-coil forming E4/K4 peptide-based fluorescence labeling. As a result, the yeast-displayed mini Q-body recognizing the anti-cancer agent methotrexate (MTX) showed significant quenching and MTX-dependent dequenching on cells. To demonstrate the applicability of the developed method to select highly responsive mini Q-bodies, a small nanobody library consisting of 30 variants that recognize human serum albumin was used as a model. The best variant, showing a 2.4-fold signal increase, was obtained through selection by flow cytometry. Furthermore, the same nanobody prepared from Escherichia coli also worked as a mini Q-body after dye labeling. The described approach will be applied to quickly obtain well-behaved Q-bodies and other fluorescent biosensors for various targets through directed evolutionary approaches

Production of an Amorphous Fe_<75>Si_<10>B_<15> Sheet by a Metallic Mold Casting Method and its Properties

The application of a metallic mold casting method to an Fe_Si_B_ alloy with the largest glass-forming ability in (Fe, Co, Ni)-Si-B system was found to cause the formation of a mostly single amorphous phase in a sheet form with a thickness of 0.1 mm. No distinct difference in thermal stability (crystallization temperature and heat of crystallization), hardness, Curie temperature and magnetization is detected between the as-cast sheet and the melt-spun amorphous ribbon with a thickness of 0.02 mm. On the other hand, bending ductility, heat of relaxation (ΔH_r) and coercivity (H_c) which can be regarded as structure sensitive properties are significantly different from those for the amorphous ribbon. That is, the sheet sample fractures during bending deformation and has a smaller ΔH_r and a larger H_c. The difference is induced presumably because the sheet sample has a more relaxed atomic configuration on a short-range scale as compared with that for the amorphous ribbon sample

Effect of Nutrient Inputs on Water Quality Change and Phytoplankton Growth in Atsumi Bay

Eutrophication in an estuary occurs as an effect of the enrichment of nutrient inputs from rivers. This condition has become one of the most common environmental issues experienced around the globe and especially in Japan. Atsumi Bay is a eutrophic coastal area in Japan. The objective of this research was to analyze the influences of nutrient inputs from the Umeda River into Atsumi Bay on pre- and post-rainfall water quality conditions. This study was conducted from July to October 2010. The results showed a decrease of surface salinity after rainfall indicating that huge freshwater inputs had overlaid the surface layer of Atsumi Bay rather than the bottom layer. Moreover, post-rainfall conditions showed an increase of chlorophyll a as an effect of phytoplankton growth, followed by an increase of particulate nutrients. On the other hand, dissolved nutrients decreased due to uptake by phytoplankton and dilution by freshwater

Assessing a Mercury Affected Area from Small-scale Gold Mining in Poboya, Central Sulawesi, Indonesia

Abstract Poboya, Central Sulawesi is one of the primary sites used for small-scale gold mining activities in Indonesia. Poboya consumes 200-500 kg of mercury/day by amalgamation. Palu, the capital city of Central Sulawesi, is a city with a population of 0.35 million people and located around 11 km away from the edge of a small-scale gold mining area in Poboya. Fifteen samples of upper layer soil and fifteen samples of plant (Calotropis gigantean) were taken along the road that connects Poboya to Palu, at every 500 m. Mercury concentration in the soil samples showed a gradual decrement as the distance from Poboya became greater. The plant samples also revealed a similar trend to the soil samples. The highest concentration of mercury in the soil was found at Poboya-i.e. 17.62 ng/mg, and the concentration of mercury in Calotropis gigantean grown at Poboya was 6.5 ng/mg. Based on the soil samples and pollution index, the data showed that the research area had heavy pollution levels up to 4 km. Moreover, the transfer factor was in the range of 0.13-3.44 at distances of 0-4 km, respectively

NMDAR2B tyrosine phosphorylation regulates anxiety-like behavior and CRF expression in the amygdala

<p>Abstract</p> <p>Background</p> <p>Anxiety disorders are a highly prevalent and disabling class of psychiatric disorders. There is growing evidence implicating the glutamate system in the pathophysiology and treatment of anxiety disorders, though the molecular mechanism by which the glutamate system regulates anxiety-like behavior remains unclear.</p> <p>Results</p> <p>In this study, we provide evidence suggesting that tyrosine phosphorylation of the NMDA receptor, an ionotropic glutamate receptor, contributes to anxiety-like behavior. The GluN2B subunit of the NMDA receptor is tyrosine-phosphorylated: Tyr-1472 is the major phosphorylation site. Homozygous knock-in mice that express a Tyr-1472-Phe mutant of GluN2B, which prevents phosphorylation of this site, show enhanced anxiety-like behavior in the elevated plus-maze test. Expression of corticotropin-releasing factor (CRF), which is important for the regulation of anxiety-like behavior, is increased in the amygdala of the knock-in mice. Furthermore, injection of CRF receptor antagonist attenuated the enhanced anxiety-like behavior of the knock-in mice. We also show that elevated plus-maze exposure simultaneously induced de-phosphorylation of Tyr-1472 and increased CRF expression.</p> <p>Conclusions</p> <p>These data suggest that Tyr-1472 phosphorylation on GluN2B is important for anxiety-like behavior by negative regulation of CRF expression in the amygdala.</p

Fetal and Neonatal Goiter in Cynomolgus Monkeys Following Administration of the Antithyroid Drug Thiamazole at High Doses to Dams During Pregnancy

To evaluate morphologic alterations in the thyroid gland in the second generation in cynomolgus monkeys, pregnant dams were exposed to high doses of thiamazole. In Experiment A, dams received thiamazole intragastrically via a nasogastric catheter from gestation day (GD) 50 to GD 150 or on the day before delivery. Initially, the dose level was 20 mg/kg/day (10 mg/kg twice daily); however, the dose level was subsequently decreased to 5 mg/kg/day (2.5 mg/kg twice daily), since deteriorated general conditions were observed in two dams. Six out of seven neonates died on the day of birth. The cause of neonatal death was tracheal compression and suffocation from goiter. The transplacental exposure to thiamazole affected the fetal thyroid glands and induced goiter in all neonates. The surviving neonate was necropsied 767 days after discontinuation of thiamazole exposure and showed reversibility of the induced changes. In Experiment B, dams were intragastrically administered thiamazole at 5 mg/kg/day (2.5 mg/kg twice daily) for treatment periods from GDs 51 to 70, 71 to 90, 91 to 110, 111 to 130 and 131 to 150. All fetuses showed enlarged thyroid glands but were viable. Histopathologically, hypertrophy and/or hyperplastic appearance of the follicular epithelium of the thyroid gland was observed at the end of each treatment period. The most active appearance of the follicular epithelium, consisting of crowded pedunculated structure, was demonstrated at end of the treatment period from GD 131 to 150. This is the first report on the morphology of fetal and neonatal goiter in the cynomolgus monkey

Change in Daily Steps and Self-efficacy of Online Interactive Exercise Classes for Community-dwelling Older Adults in Japan: A Preliminary Study

Aims: This study aimed to (1) examine the feasibility of an online interactive exercise class for community-dwelling older adults and (2) preliminarily examine changes in physical activity and self-efficacy. Methods: Participants were 25 community-dwelling older adults aged 65 years or older, but due to 5 dropouts, the final number of participants for analysis was 20 (mean age 76.9 ± 5.7 years). The intervention program was conducted for 40 minutes each session, twice a week for four consecutive weeks, using the LINE group call (LINE Corporation, Japan). An online questionnaire was used to assess participant characteristics, modified Fall Efficacy Scale score, modified Gait Efficacy Scale (m-GES) score, self-rated health, and daily steps, which were compared pre- and post-intervention using the Wilcoxon signed-rank sum and chi-square tests. Results: The Wilcoxon signed-rank sum test showed significant improvement in the m-GES score and daily steps. The chi-square test showed that self-rated health was significantly greater in the maintenance/increase group. Conclusions: Online interactive exercise classes are feasible for community-dwelling older individuals. These results also suggest the possibility of using telehealth to improve physical activity and self-efficacy