Screening of High-Affinity scFvs From a Ribosome Displayed Library Using BIAcore Biosensor

An experimental protocol was developed to screen high-affinity single-chain Fv antibody fragments (scFvs) from a Xanthomonas axonopodis pv. citri (Xac) immunized ribosome display library using BIAcore biosensor. The screening methods involved immobilizing antigen [lipopolysaccharides (LPS) of Xac] on sensor chip HPA and then unpurified expression products of scFvs flowing over the immobilized sensor chip. The affinity-improved scFvs were selected based on dissociation rate constants (k _d). Thirty-five enzyme-linked immunosorbent assay-positive scFvs were analyzed by BIAcore, and three of those (scFv A1, B2, and C5) with lower k _d were screened. To demonstrate the accuracy of the screening method, the three scFvs were expressed in Escherichia coli HB2151 and purified. The purified scFvs were subsequently further identified according to association rate and affinity constants. The results showed that the three scFvs (A1, B2, and C5) had high affinity for LPS of Xac (3.51 × 10⁻¹¹, 1.13 × 10⁻¹⁰, 5.06 × 10⁻¹⁰ M, respectively). Furthermore, the scFv B2 was highly specific for LPS of Xac and had no any cross-reactions with bovine serum albumin and LPS from Xac-related bacteria. This provided evidence that the information from the BIAcore screening assay could be accurate.     

Electrochemical sensor for heparin based on a poly(thionine) modified glassy carbon electrode

A poly(thionine) thin film modified electrode was successfully assembled on the surface of the glassy carbon electrode by means of electrochemical polymerization, which was carried out with cyclic voltammetric sweeping in the potential range 0 to +1.4 V (vs. Ag/AgCl) in perchloric acid solution containing 0.1 mmol L^?1 thionine. The film modified electrode exhibited a couple of well-defined redox peaks, and the redox peaks decreased correspondingly without a shift of the peak potential after the addition of heparin. The conditions of the binding reaction and the electrochemical detection were optimized. Under the optimum conditions the decrease of the peak current was proportional to the concentration of heparin in the range 4.0 to 22.0 μg mL^?1 and the detection limit was 0.28 μg mL^?1. The relative standard deviation (RSD) for five parallel determinations of 10.0 μg mL^?1 heparin was 0.93%. The effects of potentially interfering species were investigated and the method was successfully applied to the determination of heparin in a pharmaceutical formulation.     

Voltammetric Studies of the Interactions Between Ferrocene‐Labeled Glutathione and Proteins in Solution or Immobilized onto Surface

Glutathione (GSH) tagged with a ferrocene (Fc) label at its C‐terminal was synthesized via coupling ferrocenyl amine to glutathione using o‐(benzotriazol‐1‐yl)‐N,N,N′,N′‐tetramethyluronium (HBTU)/1‐hydroxybenzotrizole (HOBt). The presence of Fc yielded well defined voltammetric signals, rendering the Fc‐tagged GSH (GSH‐Fc) suitable for electrochemical studies of GSH binding to other biological species. The interaction of GSH‐Fc with bovine serum albumin (BSA) was investigated, and a binding ratio of 1.41±0.06 (GSH‐Fc/BSA) and an affinity constant K_a of 6.53±2.01×10⁶ M^?1 were determined. These results compare well with those measured by fluorescence using untagged GSH, suggesting that the attachment of Fc to GSH does not significantly perturb the GSH structure and binding behavior. By contrasting the binding behavior to several compounds that are known to conjugate to different domains of BSA, the voltammetric study confirmed that GSH‐Fc binds at subdomain IIA of BSA with high affinity. The versatility of GSH‐Fc for studying GSH binding to surface‐confined proteins was also demonstrated with the GSH binding to electroinactive Zn‐metallothionein (Zn₇‐MT) through hydrogen binding at the region between the Zn₇‐MT α and β domains.     

Identifying nonspecific ligand binding in electrospray ionization mass spectrometry using the reporter molecule method

The application of the reporter molecule (M_rep) method for identifying nonspecific complexes in the ES-MS analysis of protein-ligand and DNA-ligand interactions in vitro is described. To test the reliability of the method, it was applied to the ES-MS analysis of protein-carbohydrate complexes originating from specific interactions in solution and from nonspecific interactions in the ES process. These control experiments confirm the basic assumptions underlying the M_rep method, namely that nonspecific ligand binding is a random process, and that the ES droplet histories for specific and nonspecific complexes are distinct. The application of the M_rep method to the ES-MS analysis of the sequential binding of the ethidium cation, a DNA intercalator, to single and double strand oligodeoxynucleotides is also described, and highlights the general utility of the method.     

Preparative isolation of pseudolaric acids A and B,and their glucosides from the root bark of Pseudolarix kaempferi using high‐speed counter‐current chromatography

In order to provide the chemical markers for the quality control of herbal medicines, four diterpenoids, pseudolaric acids A and B (PAA and PAB), and their glucosides were isolated from the methanol extract of the Chinese herb Pseudolarix kaempferi using high‐speed counter‐current chromatography (HSCCC). The diphase solvent system was n‐hexane/EtOAc/MeOH/H₂O which was used at two ratios (5:5:5:5 and 1:9:4:6 by volume) in the separation of pseudolaric acids and their glycosides, respectively. As a result, PAA (14 mg), PAB (129 mg), PAA‐O‐β‐D ‐glucopyranoside (8 mg, PAAG), and PAB‐O‐β‐D ‐glucopyranoside (42 mg, PABG) were obtained from 0.5 g of the crude extract. Their purities were determined to be above 97% by HPLC analysis. Their chemical structures were confirmed by ¹H and ¹³C NMR analysis or HPLC comparison with the reference compounds.     

Transportation-information inequalities for Markov processes

In this paper, one investigates the transportation-information T _c I inequalities: α(T _c (ν, μ)) ≤ I (ν|μ) for all probability measures ν on a metric space ${(\mathcal{X}, d)}$ , where μ is a given probability measure, T _c (ν, μ) is the transportation cost from ν to μ with respect to the cost function c(x, y) on ${\mathcal{X}^2}$ , I(ν|μ) is the Fisher–Donsker–Varadhan information of ν with respect to μ and α : [0, ∞) → [0, ∞] is a left continuous increasing function. Using large deviation techniques, it is shown that T _c I is equivalent to some concentration inequality for the occupation measure of a μ-reversible ergodic Markov process related to I(·|μ). The tensorization property of T _c I and comparisons of T _c I with Poincaré and log-Sobolev inequalities are investigated. Several easy-to-check sufficient conditions are provided for special important cases of T _c I and several examples are worked out.     

On Extension of 1-Lipschitz Mappings between Two Unit Spheres of lp(Γ) Type Spaces (1

FANG Xi Nian 《数学研究与评论》2009,29(4)

Ionic effect on the binding of bilirubin to the imprinted poly(methacrylic acid-co-ethylene glycol dimethylacrylate)

A molecularly imprinted polymer (MIP) capable of detecting bilirubin was successfully synthesized. Bilirubin template was imprinted in poly(methacrylic acid-co-ethylene glycol dimethylacrylate) [poly(MAA-co-EGDMA)]. MAA and EGDMA were used as the monomer and the cross-linker, respectively. The optimal solvent conditions to maintain its stability were discussed. Solvent system based on ethylenediamine tetraacetic acid (EDTA) and ascorbic acid was compared with respect to the stability of bilirubin. pH and bilirubin concentration were both investigated for the bilirubin stability. Blue light as well as aeration was applied to inspect the regarding effects. The cross-linking effect was further confirmed by the thermogravimetric analysis (TGA). The effect of salts, such as NaCl and KCl on the binding capacity of the molecularly imprinted polymer was also discussed. Further, the rat serum and bile samples were applied and the binding of the MIPs for bilirubin was thus confirmed.     

Transparent conducting properties of c-axis-oriented Na_xCoO_2 epitaxial thin films

正Transparent conducting oxides(TCOs)have attracted intensive attention because of their broad application in photoelectric devices such as solar cells,light-emitting diodes,and flat-panel displays.Current TCOs used in industry,including tin(IV)oxide(SnO_2),indium oxide(In_2O_3),indium tin oxide(ITO),and zinc oxide(ZnO),are primarily n-type semiconductors[1-3].Suitable p-type TCOs are still being     

基于多光谱法的激发温度和辐射温度瞬态测试技术

近些年来,随着国内外尖端科技快速发展,温度测量无论是对于国防建设领域还是对于工业制造领域都有着极为重要的指导意义和研究价值。尤其在瞬态超高温测量方面,测温精度要求更为严苛。测温方法多种多样,多光谱法由于其精度较高且适用性强,被国内外专家广泛运用。基于多光谱测温法,提出一种新的能够同时高精度测量目标的瞬态激发温度和辐射温度的方法。该方法通过查找可信度更高的目标物理特性数据以及更为精确的多光谱直线拟合方法,精准计算得到目标激发温度。通过建立更加准确的数学模型和算法,减小光谱发射率对整个测温过程的影响实现高精度的辐射温度测量。通过相关测温实验表明,系统测温精度达到3%。