High‐Throughput Development of a Hybrid‐Type Fluorescent Glutamate Sensor for Analysis of Synaptic Transmission

Fluorescent sensors are powerful tools for visualizing cellular molecular dynamics. We present a high‐throughput screening system, designated hybrid‐type fluorescence indicator development (HyFInD), to identify optimal position‐specific fluorophore labeling in hybrid‐type sensors consisting of combinations of ligand‐binding protein mutants with small molecular fluorophores. We screened sensors for glutamate among hybrid molecules obtained by the reaction of four cysteine‐reactive fluorescence probes with a set of cysteine‐scanning mutants of the 274 amino acid S1S2 domain of AMPA‐type glutamate receptor GluA2 subunit. HyFInD identified a glutamate‐responsive probe (enhanced glutamate optical sensor: eEOS) with a dynamic range >2400 %, good photostability, and high selectivity. When eEOS was specifically tethered to neuronal surfaces, it reliably visualized the spatiotemporal dynamics of glutamate release at single synapses, revealing synapse‐to‐synapse heterogeneity of short‐term plasticity.     

Development of ambient sampling chemi/chemical ion source with dielectric barrier discharge

The development of a new configuration of chemical ionization (CI)‐based ion source is presented. The ambient air containing the gaseous sample is sniffed into an enclosed ionization chamber which is of sub‐ambient pressure, and is subsequently mixed with metastable species in front of the ion inlet of the mass spectrometer. Metastable helium atoms (He*) are used in this study as the primary ionizing agents and are generated from a dielectric barrier discharge (DBD) source. The DBD is powered by an AC high‐voltage supply and the configuration of the electrodes is in such a way that the generated plasma is confined within the discharge tube and is not extended into the ionization chamber. The construction of the ion source is simple, and volatile compounds released from the bulky sample can also be analyzed directly by approaching the sample to the sampling nozzle. When combined with heated nitrogen or other desorption methods, its application can also be extended to non‐volatile compounds, and the consumption for helium can be kept minimum solely for maintaining the stable discharge and gas phase ionization. Applications to non‐proximate sample analysis, direct determination of active ingredients in drug tablets and the detection of trace explosive such as hexamethylene triperoxide diamine are demonstrated. Copyright © 2010 John Wiley & Sons, Ltd.     

Label-free biosensors based on aptamer-modified graphene field-effect transistors

A label-free immunosensor based on an aptamer-modified graphene field-effect transistor (G-FET) is demonstrated. Immunoglobulin E (IgE) aptamers with an approximate height of 3 nm were successfully immobilized on a graphene surface, as confirmed by atomic force microscopy. The aptamer-modified G-FET showed selective electrical detection of IgE protein. From the dependence of the drain current variation on the IgE concentration, the dissociation constant was estimated to be 47 nM, indicating good affinity and the potential for G-FETs to be used in biological sensors.     

A light-controlled reversible DNA photoligation via carbazole-tethered 5-carboxyvinyluracil

We describe a light-controlled template-directed reversible DNA photoligation via carbazole-tethered 5-carboxyvinyluracil. Carbazole-tethered 5-carboxyvinyl-2'-deoxyuridine-containing oligodeoxynucleotide (ODN) can be ligated by irradiation at 366 nm in the presence of template ODN, and the ligated ODN can be split by irradiation at 366 nm in the absence of template ODN.     

Phase Equilibrium in the System Ln-Mn-O IV. Ln=Sm at 1100°C

Phase equilibrium in a Sm-Mn-O system has been established at 1100°C while changing the oxygen partial pressure from 0 to 13.00 in −log (P_O2/atm), and a phase diagram at 1100°C is presented for a Sm₂O₃-MnO-MnO₂ system. Under the experimental conditions, Sm₂O₃, MnO, Mn₃O₄, SmMnO₃, and SmMn₂O₅ phases are present at 1100°C, but Sm₂MnO₄, Mn₂O₃, and MnO₂ are unstable in the system. LnMn₂O₅- type phase is stable under the present experimental conditions differing from the previously reported La-Mn-O and Nd-Mn-O systems.A wide range of nonstoichiometry has been found in the SmMnO₃ phase which coexisted with Sm₂O₃. X ranges from −0.010 at log P_O2=−10.00 to 0.098 at log P_O2=0 in the molecular formula of SmMnO_3+X. The nonstoichiometry is represented by an equation, N_O/N_SmMnO3=3.00×10⁻⁴ (log P_O2)³ +6.20×10⁻³ (log P_O2)²+4.28×10⁻² (log P_O2)+0.0979, and the activities of the components in the solid solution are calculated using the equation. SmMnO₃ seems to vary in composition in the Sm₂O₃-rich or Sm₂O₃-poor side as it was with LaMnO₃. SmMn₂O₅ is slightly nonstoichiometric.Lattice constants of SmMnO₃ made under different oxygen partial pressures and those of SmMn₂O₅ prepared in air were determined, along with spacings and relative intensities of SmMn₂O₅. Standard Gibbs energies of reactions shown in the system were calculated and compared with previously reported values.     

Phase equilibrium in the system Y-Fe-O at 1100°C

Phase equilibrium was established in the Y-Fe-O system at 1100°C by varying the oxygen partial pressure from −log(P_O2/atm)=15.00 to 0, allowing construction of a phase diagram at 1100°C for the system Y₂O₃-Fe-Fe₂O₃. In the system, two ternary compounds, YFeO₃ and Y₃Fe₅O₁₂, were stable and had nonstoichiometric composition, whereas YFe₂O₄ was not found to be stable. The present result was different from that of previous studies at 1200°C, in which YFe₂O₄ was stable, along with the above two ternary compounds. Lattice constants of YFeO₃ and Y₃Fe₅O₁₂, prepared in air by a quenching method, were determined and compared with previous values, and showed close agreement. The standard Gibbs energy changes of the reactions in the Fe-O system, Fe+1/2O₂=FeO, 3FeO+1/2O₂=Fe₃O₄, and 2/3Fe₃O₄+1/6O₂=Fe₂O₃, were determined, and the obtained values were compared with the previous values. The standard Gibbs energy changes of the reactions, Fe+1/2Y₂O₃+3/4O₂=YFeO₃, and 3YFeO₃+2/3Fe₃O₄+1/6O₂=Y₃Fe₅O₁₂, were calculated from the oxygen partial pressures in equilibrium.     

Miscibility, mechanical and thermal properties of poly(lactic acid)/polyester-diol blends

We found new plasticizers with high molecular weight and low mobility for poly(lactic acid) (PLA). The new plasticizers are polyester-diols (PED) with nominal molecular weight 2000. Temperature dependence of oscillatory tensile moduli of PLA/PED blends in solid states was measured and miscible pairs of the blends were found. The miscible pairs are PLA/poly(ethylene adipate) and PLA/poly(diethylene adipate). Observation by scanning electron microscope and results of differential scanning calorimetry also indicate that these blends are miscible with PLA at weight ratio of PED less than or equal to 20%. In these blends, glass transition temperature is significantly lower than that of PLA. On the other hand, poly(butylene adipate) and poly(hexamethylene adipate) are partially miscible with PLA at weight ratio of PED 20%. The difference of the miscibility is discussed in terms of solubility parameter, which is calculated using the method by Small and configurational entropy.     

Atmospheric pressure chemical ionization of explosives using alternating current corona discharge ion source

The high‐sensitive detection of explosives is of great importance for social security and safety. In this work, the ion source for atmospheric pressure chemical ionization/mass spectrometry using alternating current corona discharge was newly designed for the analysis of explosives. An electromolded fine capillary with 115 µm inner diameter and 12 mm long was used for the inlet of the mass spectrometer. The flow rate of air through this capillary was 41 ml/min. Stable corona discharge could be maintained with the position of the discharge needle tip as close as 1 mm to the inlet capillary without causing the arc discharge. Explosives dissolved in 0.5 µl methanol were injected to the ion source. The limits of detection for five explosives with 50 pg or lower were achieved. In the ion/molecule reactions of trinitrotoluene (TNT), the discharge products of NO_x^? (x = 2,3), O₃ and HNO₃ originating from plasma‐excited air were suggested to contribute to the formation of [TNT ? H]^? (m/z 226), [TNT ? NO]^? (m/z 197) and [TNT ? NO + HNO₃]^? (m/z 260), respectively. Formation processes of these ions were traced by density functional theory calculations. Copyright © 2015 John Wiley & Sons, Ltd.     

In situ analysis of soybeans and nuts by probe electrospray ionization mass spectrometry

The probe electrospray ionization (PESI) is an ESI‐based ionization technique that generates electrospray from the tip of a solid metal needle. In the present work, we describe the PESI mass spectra obtained by in situ measurement of soybeans and several nuts (peanuts, walnuts, cashew nuts, macadamia nuts and almonds) using different solid needles as sampling probes. It was found that PESI‐MS is a valuable approach for in situ lipid analysis of these seeds. The phospholipid and triacylglycerol PESI spectra of different nuts and soybean were compared by principal component analysis (PCA). PCA shows significant differences among the data of each family of seeds. Methanolic extracts of nuts and soybean were exposed to air and sunlight for several days. PESI mass spectra were recorded before and after the treatment. Along the aging of the oil (rancidification), the formation of oxidated species with variable number of hydroperoxide groups could be observed in the PESI spectra. The relative intensity of oxidated triacylglycerols signals increased with days of exposition. Monitoring sensitivity of PESI‐MS was high. This method provides a fast, simple and sensitive technique for the analysis (detection and characterization) of lipids in seed tissue and degree of oxidation of the oil samples. Copyright © 2015 John Wiley & Sons, Ltd.     

Desorption Mass Spectrometry for Nonvolatile Compounds Using an Ultrasonic Cutter

In this work, desorption of nonvolatile analytes induced by friction was studied. The nonvolatile compounds deposited on the perfluoroalkoxy substrate were gently touched by an ultrasonic cutter oscillating with a frequency of 40 kHz. The desorbed molecules were ionized by a dielectric barrier discharge (DBD) ion source. Efficient desorption of samples such as drugs, pharmaceuticals, amino acids, and explosives was observed. The limits of detection for these compounds were about 1 ng. Many compounds were detected in their protonated forms without undergoing significant fragmentation. When the DBD was off, no ions for the neutral samples could be detected, meaning that only desorption along with little ionization took place by the present technique.