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PubMed2005

Spectroscopic identification of S-Au interaction in cysteine capped gold nanoparticles.

This study deals with the synthesis of cysteine capped gold nanoparticles with an average size of 12 nm by borohydride reduction and spectroscopic identification of SAu interaction. We have studied the interaction of thiol with gold nanoparticles in aqueous medium by employing UV-vis, Raman, NMR, and FT-IR spectroscopy. The shifting of gold plasmon resonance in the UV-vis spectra shows the stabilization of gold nanoparticles by cysteine. The disappearance of S-H stretching in both the IR and Raman spectra and the shifting of the NMR signals of the protons in close proximity to the metal center supported the existence of the S-Au interaction in cysteine capped gold nanoparticles. The TEM images shows cysteine capped gold nanoparticles as distinct and spherical entities as compared to free colloidal gold nanoparticles.

Cysteine↗
PubMed2005

Cytotoxicity of nanosize V(2)O(5) particles to selected fibroblast and tumor cells.

Two kinds of nanosize V(2)O(5) particles were synthesized in our own laboratory and concomitantly applied to V79 and L929 fibroblasts and SCCVII, B16F10 and FsaR tumor cells. The morphologies of the cells were monitored using an inverted inverse microscope equipped with digital camera, while quantitative determination of the cytotoxicity of nanosize V(2)O(5) particles was measured using crystal violet bioassay. Twenty four hours after the addition of nanosize V(2)O(5) particles (20muM), noticeable changes in the morphology and density of fibroblast and cancer cells were observed. Reculturing in a freshly prepared medium for the next 24h showed a high recovery effect on V79, SCCVII and B16F10 cells, while FsaR and L929 cells were seriously damaged and unable to recover. At a higher concentration of nanosize V(2)O(5) particles (100muM), the cytotoxicity of V(2)O(5) prevailed against the recovery effect in all cell types. Quantitative measurements have shown that the resistance of investigated cell cultures to the cytotoxicity of nanosize V(2)O(5) particles decreases in the order V79>SCCVII>B16F10>FsaR>L929. The high cytotoxic effect found on FsaR cells suggests that nanosize V(2)O(5) particles could be regarded as poisoning material in the treatment of FsaR fibrosarcoma cells. Possible mechanisms involved in the cytotoxicity of nanosize V(2)O(5) particles were discussed.

Cell Line↗
PubMed1995

Calcium pump kinetics determined in single erythrocyte ghosts by microphotolysis and confocal imaging.

The activity of the plasma membrane calcium pump was measured in single cells. Human red blood cell ghosts were loaded with a fluorescent calcium indicator and either caged calcium and ATP (protocol A) or caged ATP and calcium (protocol B). In a suitably modified laser scanning microscope either calcium or ATP were released by a short UV light pulse. The time-dependent fluorescence intensity of the calcium indicator was then followed in single ghosts by repetitive confocal imaging. The fluorescence intensity was converted into calcium concentration, which in turn was used to derive the kinetic parameters of the calcium pump, the Michaelis-Menten constant Km, and the maximal transport rate vmax. Km and vmax values derived in this manner were 24 +/- 14 microM and 1.0 +/- 0.6 microM/(ghost s) for protocol A, and 4 +/- 3 microM and 1.0 +/- 0.6 microM/(ghost s) for protocol B, respectively. The difference between A and B is presumably caused by calmodulin, which is inactive in the experiments with protocol A. The possibilities to extend the new method to living nucleus-containing cells transiently transfected with mutants of the plasma membrane calcium pump are discussed.

Adenosine Triphosphate↗
PubMed1987

Characterization of organotin species using microbore and capillary liquid chromatographic techniques with an epifluorescence microscope as a novel imaging detector.

The novel application of a UV epifluorescence microscope as an imaging detector for microbore and capillary high-performance liquid chromatography (HPLC) is reported. The microscope is focused on an in-line quartz flow cell incorporated down stream of a microbore HPLC column or directly on an optically clear portion of fused-silica capillary columns for analyte detection. The effect of different fluorescent ligand to analyte ratios on detection limits is also reported, as well as the effect of different image volume sizes produced by changes in microscope objective lens magnification power. Determination of relative sensitivities an detection limits for methyl- and butyltin compounds, complexed with fluorescent dyes, reveals that the organotins show decreasing sensitivity as the number of alkyl substituents on the tin atom increases, with minimum detectable amounts of 6-160 pg of analyte-ligand complex.

Chromatography, High Pressure Liquid↗
PubMed2001

Poly(glycidyl methacrylate-divinylbenzene-triallylisocyanurate) continuous-bed protein chromatography.

A novel continuous bed with high dynamic adsorption capacity for protein has been developed. It is a macroporous poly(glycidyl methacrylate-divinylbenzene-triallylisocyanurate) rod prepared by in situ copolymerization in a chromatographic tube. The bed matrix contained epoxy groups, so diethylaminohydroxypropyl groups were coupled to the matrix, leading to an anion-exchange continuous bed. The component, specific surface area, and the pore structure of the bed matrix were characterized by Fourier transform infrared spectroscopy, BET method and scanning and transmission electron microscopies, respectively. The flow properties, column efficiency and the dynamic adsorption behavior of the bed were studied. The results show that the continuous bed, a ternary copolymer of glycidyl methacrylate (GMA), divinylbenzene (DVB) and triallylisocyanurate (TAIC) with a specific surface area of 56.4 m2/g, consisted of a three-dimensional structure made up of continuous clusters of microspheres (300 nm) and interconnected irregular pores. The rate of mass transfer is enhanced by the convection of the mobile phase through the pores. The dynamic adsorption isotherm of the anion-exchange column for bovine serum albumin was expressed by the Langmuir equation with a dynamic capacity as high as 76.0 mg/g. Moreover, the separation of proteins, i.e. lysozyme, hemoglobin and bovine serum albumin, is little affected by mobile-phase velocity up to 902.5 cm/h; it was completed within 5 min at 902.5 cm/h.

Adsorption↗
PubMed2001

Photopolymerized sol-gel frits for packed columns in capillary electrochromatography.

Porous sol-gel frits are fabricated in a capillary column by filling it with a solution of 3-(trimethoxysilyl)propyl methacrylate, hydrochloric acid, water, toluene (porogen), and a photoinitiator (Irgacure 1800) and exposing it to UV light at 365 nm for 5 min. The separation column (30 cm x 75 microm I.D.) contains between the inlet and outlet frits a 15-cm packed segment filled with 5-microm silica particles modified with the chiral compound (S)-N-3,5-dinitrobenzoyl-1-naphthylglycine. A detection window (1 mm long) is placed immediately after the outlet frit. To demonstrate the performance of this chiral separation column, mixtures of 16 different amino acids (three of which are not naturally occurring) derivatized with the fluorogenic reagent 4-fluoro-7-nitro-2,1,3-benzoxadiazole were separated by capillary chromatography. The enantiomeric separation of the column results in a resolution ranging from 1.21 to 8.29, and a plate height ranging from 8.7 to 39 microm.

Chromatography, Micellar Electrokinetic Capillary↗
PubMed1997

Photo-immobilization of epidermal growth factor enhances its mitogenic effect by artificial juxtacrine signaling.

Photo-reactive epidermal growth factor (EGF) was synthesized by coupling EGF with azidobenzoic acid and was immobilized onto the wells of a polystyrene culture plate by photo-irradiation. The photo-immobilized EGF enhanced the growth of anchorage-dependent cells more than native or azidobenzoyl derivatized EGF. A small amount of photo-immobilized EGF was sufficient to enhance the growth of cells and the maximal mitogenic effect was greater than that of native or derivatized EGF. On the other hand, the photo-immobilized EGF did not enhance growth of anchorage-independent cells. In addition, signal transduction in the cells adhered only on the EGF-immobilized surface was observed by staining of phosphotyrosine residues by anti-phosphotyrosine antibodies. These results showed that the enhanced cell growth was due to direct interaction between the cells and the immobilized EGF. Photo-immobilization could be a universal means of fixing growth factors onto an artificial matrix that is devoid of chemically functional groups scaffolding growth factors and could provide a new tool to elucidate signal transduction mechanism and could lead to the development of a new protein-free cell culture system or tissue engineering materials.

Animals↗
PubMed2000

Chemoenzymatical grafting of acrylamide onto lignin.

Laccase (E.C.1.10.3.2.) from white-rot basidiomycete Trametes versicolor and dioxane peroxides were essential in the copolymerization of acrylamide and a derivative with lignin in a dioxane-H(2)O (7:3) mixture. Both a solubility test and an elemental analysis of the eluted and separated fractions provided evidence of grafting. Gel permeation chromatography on Sephadex G-100 showed that the side chain and the lignin backbone migrate as one unit through the column. Copolymers obtained were characterized by UV-VIS-spectroscopy as well as FT-IR, and 13C-CPMAS-NMR spectroscopy. Freeze-dried copolymers of lignin and acrylamide appeared as homogeneous fibril-like particulate. The mechanism of the enzymatical grafting is discussed.

Acrylamide↗
PubMed2000

The effect of liquid crystalline structure on chlorhexidine diacetate release.

The aim of this study was to examine different liquid crystalline preparations containing chlorhexidine diacetate and to find connection between their structure and the kinetic of drug release. Nonionic surfactant, Synperonic A7 (PEG(7)-C(13-15)) was selected for the preparation of the examined liquid crystalline systems. Mixtures of different ratios of Synperonic A7 and water were produced. By increasing the water content of the systems, lamellar and hexagonal liquid crystal structures were observed. For the analysis of the prepared liquid crystalline systems polarising microscopy, rheology study, differential scanning calorimetry and dynamic swelling tests were carried out. The chlorhexidine diacetate release was examined by Franz-type vertical diffusion cell apparatus. The chlorhexidine diacetate release from hexagonal liquid crystalline preparations was characterised by zero-order release kinetics, while the drug release from lamellar liquid crystalline systems was described by anomalous (non-Fickian) transport. The results indicate that the drug release kinetic is strongly dependent on the liquid crystalline structure.

Anti-Infective Agents, Local↗
PubMed1979

Structure of beta-casein.

The temperature and concentration dependent association of beta-casein was studied by means of viscometry, gel filtration chromatography, electron microscopy, analytical ultracentrifugation and UV difference spectrophotometry. Degrees of polymerization of 12, 22 and 49 and free energies of association of -21, -23 and -25kJ/mole monomer were found at temperatures of 10, 15 and 20 degrees C respectively in 0.2 M Na phosphate buffer pH 6.7. Monomeric beta-casein was not a completely random coil but became more compact with increasing temperature, due to hydrophobic interactions.

Calorimetry↗
PubMed1991

Medical technology transfer in major Chinese medical schools.

This paper examines how the decision-making process and its consequences affect medical technology transfer in major Chinese medical schools. Data are from a 1987 survey of 13 key medical universities, directly supervised by the Ministry of Public Health in the People's Republic of China. This paper limits itself to four types of laboratory equipment--electron microscopes, UV/VIS spectrophotometers, high-performance liquid chromatographs, and polygraphs. Decisions on the transfer of medical technology have been more decentralized in China since the economic reform in 1978. The major reason for schools to import these four types of equipment is their dissatisfaction with the quality of domestic products. Chinese medical schools depend heavily on the information provided at medical equipment exhibits and their neighboring schools. Their decisions to acquire the equipment are based more on the quality and service available than on the prices. Chinese medical schools face serious infrastructure problems in acquiring and maintaining these pieces of equipment. A number of suggestions are made for improving the efficiency of medical technology transfer in China.

China↗
PubMed2000

Segregation of micrometer-dimension biosensor elements on a variety of substrate surfaces.

With the rapid development of micro total analysis systems and sensitive biosensing technologies, it is often desirable to immobilize biomolecules to small areas of surfaces other than silicon. To this end, photolithographic techniques were used to derivatize micrometer-sized, spatially segregated biosensing elements on several different substrate surfaces. Both an interference pattern and a dynamic confocal patterning apparatus were used to control the dimensions and positions of immobilized regions. In both of these methods, a UV laser was used to initiate attachment of a photoactive biotin molecule to the substrate surfaces. Once biotin was attached to a substrate, biotin/avidin/biotin chemistry was used to attach fluorescently labeled or nonlabeled avidin and biotinylated sensing elements such as biotinylated antibodies. Dimensions of 2-10 microm were achievable with these methods. A wide variety of materials, including glassy carbon, quartz, acrylic, polystyrene, acetonitrile-butadiene-styrene, polycarbonate, and poly(dimethylsiloxane), were used as substrates. Nitrene- and carbene-generating photolinkers were investigated to achieve the most homogeneous films. These techniques were applied to create a prototype microfluidic sensor device that was used to separate fluorescently labeled secondary antibodies.

Biosensing Techniques↗
PubMed1991

Wild-type and mutant bacteriorhodopsins D85N, D96N, and R82Q: purification to homogeneity, pH dependence of pumping, and electron diffraction.

Bacterioopsin, expressed in Escherichia coli as a fusion protein with 13 heterologous residues at the amino terminus, has been purified in the presence of detergents and retinylated to give bacteriorhodopsin. Further purification yielded pure bacteriorhodopsin, which had an absorbance ratio (A280/A lambda max) of 1.5 in the dark-adapted state in a single-detergent environment. This protein has a folding rate, absorbance spectrum, and light-induced proton pumping activity identical with those of bacteriorhodopsin purified from Halobacterium halobium. Protein expressed from the mutants D85N, D96N, and R82Q and purified similarly yielded pure protein with absorbance ratios of 1.5. Proton pumping rates of bacteriorhodopsins with the wild-type sequence and variants D85N, D96N, and R82Q were determined in phospholipid vesicles as a function of pH. D85N was inactive at all pH values, whereas D96N was inactive from pH 7.0 to pH 8.0, where wild type is most active, but had some activity at low pH. R82Q showed diminished proton pumping with the same pH dependence as for wild type. Bacteriorhodopsin purified from E. coli crystallized in two types of two-dimensional crystal lattices suitable for low-dose electron diffraction, which permit detailed analysis of structural differences in site-directed variants. One lattice was trigonal, as in purple membrane, and showed a high-resolution electron diffraction pattern from glucose-sustained patches. The other lattice was previously uncharacterized with unit cell dimensions a = 127 A, b = 67 A, and symmetry of the orthorhombic plane group pgg.

Bacteriorhodopsins↗
PubMedDate not supplied

Microcapsule modification with peroxidase-catalyzed phenol polymerization.

A biocatalytic polymer synthesis on a surface of polyelectrolyte microcapsules was studied. Horseradish peroxidase assembled in nanoorganized capsule walls by alternate adsorption with linear polyions retains its activity in reactions of enzyme-catalyzed polymerization of 4-oxyphenols. It allowed controllable synthesis of a phenolic polymer layer on microcapsule walls using an outermost surface peroxidase layer as a template. By varying the phenol type, buffer pH, and reaction component concentrations, the phenolic polymer coating of the capsules with a thickness in the range 20-50 nm was formed. The polymeric products are fluorescent, which provided a good opportunity for confocal image analysis of the capsule wall structure and the attached layer. The influence of a phenolic polymer layer on the permeability of the capsule walls was investigated.

Catalysis↗
PubMedDate not supplied

Protein adsorption modalities on polyelectrolyte multilayers.

Protein adsorption on polyelectrolyte multilayers (PEMUs) was evaluated using a combination of synthetic polyelectrolytes and proteins, including serum albumin, fibrinogen, and lysozyme. Variables such as surface and protein charge, polymer hydrophobicity, and hydrophilic repulsion were introduced to probe interaction mechanisms. Quantitative analysis with reflectance Fourier transform infrared spectroscopy, optical waveguiding, and UV-vis absorption, together with qualitative information from atomic force microscopy, provided a coordinated picture for what drives protein adsorption and how the molecules are disposed on the multilayer surface. It was found that multilayers bearing a particular surface charge sorbed biomolecules if they were of opposite charge, yielding significant loadings within the bulk PEMU. Adsorption of like-charged proteins, as surface aggregates, occurred to a much lower extent, driven by nonelectrostatic forces. A diblock copolymer comprising a hydrophilic poly(ethylene oxide) block was capable of further minimizing protein adsorption as a result of hydrophilic repulsion, although none of the surfaces tested defeated protein adsorption completely. However, poly(acrylic acid) homopolymer was quite effective in this respect. A composition gradient, formed during multilayer buildup, induced a gradient in hydrophilicity through the PEMU, which is an efficient and economical method of creating a protein-resistant surface.

Adsorption↗
PubMedDate not supplied

Physicochemical properties and structural changes in vegetative tissues as affected by a direct current electrical field.

Cylindrical pieces of potato, sweet potato, kohlrabi, radish, and pear were interposed between a pair of electrodes, and a direct current was applied. A special custom-made apparatus enabled the use of differently shaped electrodes. The electrical field was applied for 1 min at 40 V/cm and caused a reduction in specimen weight by a minimal value of 2.7% of initial weight in sweet potato to a maximum 38.4% in pear. The affected area of the tissue resembled the shape of the electrode. Pores were produced in the tissue (from the anode side), possibly promoting slow release of minerals and other cell components from the contracted specimens. From the cathode side, cell "sealing" could be observed. Weight loss was dependent on the mechanical properties of the nontreated vegetative tissue specimens. After confirmation that all samples pass through induced electrical shrinkage, further work, executed only on potato, demonstrated that after electrical treatment the samples were less brown (higher L values). In addition, a dependence of weight loss on current intensity, electrode diameter, and surface ratio between the electrode and specimen was shown. The reduction in weight loss could be useful for initial drying of vegetative materials. Indirect proof of a decrease in enzyme activity as a result of electrical field application could be beneficial in replacing traditional methods for browning prevention.

Catechol Oxidase↗