Rapid microchip-based electrophoretic immunoassays for the detection of swine influenza virus

Towards developing rapid and portable diagnostics for detecting zoonotic diseases, we have developed microchip-based electrophoretic immunoassays for sensitive and rapid detection of viruses. Two types of microchip-based electrophoretic immunoassays were developed. The initial assay used open channel electrophoresis and laser-induced fluorescence detection with a labeled antibody to detect influenza virus. However, this assay did not have adequate sensitivity to detect viruses at relevant concentrations for diagnostic applications. Hence, a novel assay was developed that allows simultaneous concentration and detection of viruses using a microfluidic chip with an integrated nanoporous membrane. The size-exclusion properties of the in situ polymerized polyacrylamide membrane are exploited to simultaneously concentrate viral particles and separate the virus/fluorescent antibody complex from the unbound antibody. The assay is performed in two simple steps--addition of fluorescently labeled antibodies to the sample, followed by concentration of antibody-virus complexes on a porous membrane. Excess antibodies are removed by electrophoresis through the membrane and the complex is then detected downstream of the membrane. This new assay detected inactivated swine influenza virus at a concentration four times lower than that of the open-channel electrophoresis assay. The total assay time, including device regeneration, is six minutes and requires <50 microl of sample. The filtration effect of the polymer membrane eliminates the need for washing, commonly required with surface-based immunoassays, increasing the speed of the assay. This assay is intended to form the core of a portable device for the diagnosis of high-consequence animal pathogens such as foot-and-mouth disease. The electrophoretic immunoassay format is rapid and simple while providing the necessary sensitivity for diagnosis of the illness state. This would allow the development of a portable, cost-effective, on-site diagnostic system for rapid screening of large populations of livestock, including sheep, pigs, cattle, and potentially birds.     

Electrochemical and spectroscopic behavior of iron(III) porphyrazines in Langmuir-Schafer films

Thin films of a newly synthesized iron(III) porphyrazine, LFeOESPz ( L = ClEtO, OESPz = ethylsulfanylporphyrazine), have been deposited by the Langmuir-Schafer (LS) technique (horizontal lifting) on ITO or gold substrates. Before deposition, the floating films have been investigated at the air-water interface by pressure/area per molecule (pi/ A) experiments, Brewster angle microscopy (BAM) and UV-vis reflection spectroscopy (RefSpec). The complex reacts with water subphase (pH 6.2) forming the mu-oxo dimer, which becomes the predominant component of the LS films ( LS-Fe) as indicated by optical, IR, XPS, and electrochemical data. LS-Fe multilayers exhibit, between open circuit potential (OCP) and +0.90 V (vs SCE), two independent peak pairs with formal potentials, E surf (I) and E surf(II) of +0.56 V and +0.78 V, respectively. According to dynamic voltammetric and coulometric experiments the peak pair at +0.56 V is attributed to one-electron process at the iron(III) centers on the monomer, while the peak pair at +0.78 V is associated to a four-electron process involving mu-oxo-dimer oligomers. LS-Fe films prove to be quite stable electrochemically between OCP and +0.90 V. The electrochemical stability decreases, however, when the potential range is extended both anodically and cathodically outside these limits, due to formation of new species. Upon incubation with TCA solutions, LS-Fe films show remarkable changes in the UV-vis spectra, which are consistent with a significant mu-oxo dimer --> monomer conversion. Addition of TCA to the electrochemical cell using a LS-Fe film as working electrode, results in a linear increase of a cathodic current peak near -0.40 V as the TCA concentration varies in the 0.1-2.0 mM range. This behavior is interpreted in terms of TCA inducing a progressive change in the composition of the LS-Fe films in favor of the monomeric iron(III) porphyrazine, which is responsible for the observed increase in the cathodic current near -0.40 V.     

Decorated DNA-Based Scaffolds as Lateral Flow Biosensors

Here we develop Lateral Flow Assays (LFAs) that employ as functional elements DNA-based structures decorated with reporter tags and recognition elements. We have rationally re-engineered tile-based DNA tubular structures that can act as scaffolds and can be decorated with recognition elements of different nature (i.e. antigens, aptamers or proteins) and with orthogonal fluorescent dyes. As a proof-of-principle we have developed sandwich and competitive multiplex lateral flow platforms for the detection of several targets, ranging from small molecules (digoxigenin, Dig and dinitrophenol, DNP), to antibodies (Anti-Dig, Anti-DNP and Anti-MUC1/EGFR bispecific antibodies) and proteins (thrombin). Coupling the advantages of functional DNA-based scaffolds together with the simplicity of LFAs, our approach offers the opportunity to detect a wide range of targets with nanomolar sensitivity and high specificity.     

Doping dependence of the pressure response of Tc in the SmO(1-x)F(x)FeAs superconductors

The superconducting transition temperature of the high-Tc SmO1-xFxFeAs superconductors increases monotonically as the F-doping level x increases to 0.20. High-pressure magnetization experiments reveal a strong sensitivity of Tc to interatomic distances in the underdoped regime (x 相似文献   

(S)-(-)-alpha,alpha-Di(2-naphthyl)-2-pyrrolidinemethanol, a useful tool to study the recognition mechanism in chiral ligand-exchange chromatography

A dynamic coating of the RP-18 carbon chain layers with the new chiral selector (S)-(-)-alpha,alpha-di(2-naphthyl)-2-pyrrolidinemethanol allowed the formation of a mixed chiral stationary phase that has been used in the separation of a selected set of amino acid racemates. Both a representative model and classification structure-property relationship studies have been performed in order to study the contribution of hydrophobic, bulky and electron-donating groups in the side chain of the chiral selector to the mechanism of chiral recognition.     

Investigation into the reactivity of the coordinatively unsaturated phosphonodithioato [Ni(MeOpdt)2] towards 2,4,6-tris(2-pyridyl)-1,3,5-triazine: goals and achievements

The reaction between the coordinatively unsaturated phosphonodithioato complex [Ni(MeOpdt)2] (1) [MeOpdt = (MeO)(4-MeOC(6)H(4))PS2-] and tptz [2,4,6-tris(2-pyridyl)-1,3,5-triazine] has been investigated. Spectrophotometric and conductometric titrations showed the formation of a neutral and an ionic species, i.e. [Ni(MeOdtp)2(tptz)] (2) and [Ni(tptz)2](MeOdtp)2 (3), in correspondence to 1 : 1 and 2 : 1 tptz : ratios, respectively. XRD studies confirmed the formation of both complexes isolated in the compounds 2.MeOH and 3.4H(2)O. In the neutral complex 2 the central Ni(II) ion features a distorted octahedral coordination, achieved through three N-atoms of tptz and three S-atoms belonging to two MeOpdt anions, one of which unexpectedly acts as a monodentate S-donor. In 3.4H(2)O, the two phosphonodithioato anions are non-coordinating and counterbalance the charge of the [Ni(tptz)2](2+) distorted octahedral complex. From the reaction 2 of with I2 and Br2, crystals of [Ni(tptz)2](I3)2 (5) and [Ni(tptz)Br(micro-Br)]2 (6) have been obtained. The dinuclear complex 6 features a structure showing tubular canals with openings of about 6 x 6 A.     

Development of a CZE method for the determination of mizolastine and its impurities in pharmaceutical preparations using response surface methodology

A fast and selective CZE method for the determination of mizolastine and related impurities is described. Response surface methodology was applied to study the influence of phosphate/triethanolamine (TEA) buffer concentration, heptakis(2,3,6-tri-O-methyl)-beta-CD (TMbetaCD) concentration, voltage and temperature. The optimum conditions were: 105 mM phosphate/TEA buffer (pH 3.0) containing 10 mM TMbetaCD, temperature 19 degrees C and voltage 30 kV. Validation of the method was performed in drug substance and drug product. Robustness was evaluated using a Plackett-Burman design, including pH among the considered factors. Applying the optimal conditions, the nine peaks were baseline separated in about 10 min. The method was applied to the quality control of mizolastine in controlled-release tablets.     

Electronic structure of neutral and monoanionic tris(benzene-1,2-dithiolato)metal complexes of molybdenum and tungsten

The reaction of 3 equiv of the ligand 2-mercapto-3,5-di-tert-butylaniline, H2[LN,S], or 3,5-di-tert-butyl-1,2-benzenedithiol, H2[LS,S], with 1 equiv of [MoO2(acac)2] or WCl6 (acac=acetonylacetate(1-)) in methanol or CCl4 afforded the diamagnetic neutral complexes [MoV(LN,S)2(L*N,S)]0 (1), [MoV(LS,S)2(L*S,S)] (2), and [WV(LS,S)2(L*S,S)] (3), where (L*N,S)- and (L*S,S)- represent monoanionic pi-radical ligands (Srad=1/2), which are the one-electron oxidized forms of the corresponding closed-shell dianions (LN,S)2- and (LS,S)2-. Complexes 1-3 are trigonal-prismatic members of the electron-transfer series [ML3]z (z=0, 1-, 2-). Reaction of 2 and 3 with [N(n-Bu)4](SH) in CH2Cl2 under anaerobic conditions afforded paramagnetic crystalline [N(n-Bu)4][MoV(LS,S)3] (4) and [N(n-Bu)4][WV(LS,S)3] (5). Complexes 1-5 have been characterized by X-ray crystallography. S K-edge X-ray absorption and infrared spectroscopy prove that a pi-radical ligand (L*S,S)- is present in neutral 2 and 3, whereas the monoanions [MV(LS,S)3]- contain only closed-shell dianionic ligands. These neutral species have previously been incorrectly described as [MVI(L)3]0 complexes with a MoVI or WVI (d0) central metal ion; they are, in fact MV (d1) (M=Mo, W) species: [MoV(LS,S)2(L*S,S)] and [WV(LS,S)2(L*S,S)] with a diamagnetic ground state St=0, which is generated by intramolecular, antiferromagnetic coupling between the MV (d1) central ion (SM=1/2) and a ligand pi radical (L*S,S)- (Srad=1/2).     

Electronic structures of tris(dioxolene)chromium and tris(dithiolene)chromium complexes of the electron-transfer series [Cr(dioxolene)(3)](z) and [Cr(dithiolene)(3)](z) (z = 0, 1-, 2-, 3-). A combined experimental and density functional theoretical study

From the reaction mixture of 3,6-di-tert-butylcatechol, H2[3,6L(cat)], [CrCl3(thf)3], and NEt3 in CH3CN in the presence of air, the neutral complex [CrIII(3,6L*(sq))3] (S = 0) (1) was isolated. Reduction of 1 with [Co(Cp)2] in CH2Cl2 yielded microcrystals of [Co(Cp)2][CrIII(3,6L*(sq))2(3,6L(cat))] (S = 1/2) (2) where (3,6L*(sq)(1-) is the pi-radical monoanionic o-semiquinonate of the catecholate dianion (3,6Lcat)(2-). Electrochemistry demonstrated that both species are members of the electron-transfer series [Cr(3,6LO,O)]z (z = 0, 1-, 2-, 3-). The corresponding tris(benzo-1,2-dithiolato)chromium complex [N(n-Bu)4][CrIII(3,5L*S,S)2(3,5LS,S)] (S = 1/2) (3) has also been isolated; (3,5LS,S)(2-) represents the closed-shell dianion 3,5-di-tert-butylbenzene-1,2-dithiolate(2-), and (3,5L*S,S)(1-) is its monoanionic pi radical. Complex 3 is a member of the electron-transfer series [Cr(3,5L(S,S))3]z (z = 0, 1-, 2-, 3-). It is shown by Cr K-edge and S K-edge X-ray absorption, UV-vis, and EPR spectroscopies, as well as X-ray crystallography, of 1 and 3 that the oxidation state of the central Cr ion in each member of both electron-transfer series remains the same (+III) and that all redox processes are ligand-based. These experimental results have been corroborated by broken symmetry density functional theoretical calculations by using the B3LYP functional.