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At least 541 records · Page 30Linked to original sources
PubMed2006

Resonance Rayleigh scattering study of interaction of hyaluronic acid with ethyl violet dye and its analytical application.

In near weak acid to neutral medium, ethyl violet (EV) can react rapidly with hyaluronic acid (HA) to form a complex, which results in a significant enhancement of resonance Rayleigh scattering (RRS) and an appearance of a new spectrum, and the scattering wavelengths appear at 231, 274, 326, 498 and 640 nm. The maximum scattering wavelength appears at 326 nm. The RRS intensity is directly proportional to the concentration of HA in the range of 0.4-48.0 microg mL(-1). A new method for the determination of trace amounts of HA based on the RRS method has been developed. The method exhibits high sensitivity, and the detection limit for HA is 9.6 x 10(-2) microg mL(-1). This method was applied for determining HA in eyedrops and in sodium hyaluronate injection samples with satisfactory results. Furthermore, the enhancement reasons of RRS and the relationship between RRS spectral characteristics of the HA-EV complex and its absorption spectrum have been discussed.

Biosensing Techniques↗
PubMed2005

Waveguide excitation fluorescence microscopy: a new tool for sensing and imaging the biointerface.

A novel biosensing and imaging technique, the waveguide excitation fluorescence microscope, has been developed for the dynamic and quantitative investigation of bio-interfacial events in situ, ranging from ligand-receptor binding to focal adhesion formation in cell-surface interactions. The technique makes use of the evanescent field created when light travels in a mono-mode, planar optical waveguide to excite fluorescence in the near interface region. Advantages of the technique include high target sensitivity for fluorescence detection (femtomolar range), high surface specificity (ca. 100 nm perpendicular to the waveguide), large area analysis with submicron resolution, 'built-in' calibration of fluorescent light gain, and the capability to perform multi-colour imaging in situ and in real time. In this work, the sensitivity of the system has already been demonstrated through dynamic measurements of the streptavidin-biotin binding event to below 20 pM concentrations, signal to noise comparisons with conventional fluorescence microscopy have shown more than a 10-fold improvement, and surface specificity of the technique has also been illustrated in a comparison of fibroblast focal adhesion images. Thus, this new tool can be used to illuminate processes occurring at the interface between biology and synthetic surfaces in a unique manner.

Biosensing Techniques↗
PubMed2005

Label free analysis of transcription factors using microcantilever arrays.

We report the measurement of protein interaction with double-stranded DNA oligonucleotides using cantilever microarray technology. We investigated two different DNA-binding proteins, the transcription factors SP1 and NF-kappaB, using cantilever arrays as they allow label-free measurement of different biomolecular interactions in parallel. Double-stranded DNA oligonucleotides containing a specific binding site for a transcription factor were sensitized on gold-coated cantilevers. The binding of the transcription factor creates a surface stress, resulting in a bending of the cantilevers. Both transcription factors could be detected independently at concentrations of 80-100 nM. A concentration dependence of the bending signal was measured using concentrations from 100 to 400 nM of NF-kappaB. The experiments show that the recognition sequence of one transcription factor can serve as a reference for the other, highlighting the sequence specificity of transcription factor binding.

DNA↗
PubMed2005

A proteomic biosensor for enteropathogenic E. coli.

The study of proteins and the molecules with which they interact on an organismwide scale is critical to understanding basic biology, and understanding and improving human health. New platform technologies allowing label-free, quantitative array-based analysis of proteins are particularly desirable. We have developed an analytical technology, reflective interferometry (RI), which provides specific, rapid, and label-free optical detection of biomolecules in complex mixtures. In order to evaluate the suitability of RI for proteomics, we have prepared a series of arrays bearing the extracellular domain of the secreted enteropathogenic Escherichia coli (EPEC) protein Translocated Intimin Receptor (Tir). These arrays are able to selectively detect the extracellular domain of the protein Intimin, Tir's natural binding partner. Furthermore, we demonstrate the use of RI and Tir-functionalized arrays for the selective detection of EPEC directly from culture.

Animals↗
PubMed2005

Measurement of the optical parameters of purple membrane and plant light-harvesting complex films with optical waveguide lightmode spectroscopy.

Purple membrane (bacteriorhodopsin) and plant light-harvesting complexes (LHCII) were dried on the optical waveguide sensor with varying thicknesses in a wide range (from 20 to several hundreds of nanometers) and the optical parameters were studied with optical waveguide lightmode spectroscopy. It was found that applying the approximate 4-layer mode equations for the measured effective refractive indices resulted in unacceptable results for the optical parameters: with increasing thickness the refractive index decreased monotonously from 1.5 to 1.1. Therefore an inverse waveguide numerical method was developed and used to obtain reliable results from the experiments. The inverse method yielded an approximately constant (1.53) refractive index independently of the thickness for the purple membrane and LHCII films. Light-induced changes in the optical parameters of the purple membrane and LHCII films were also studied. For purple membrane films the most significant effect is the change in refractive index and absorption. For LHCII films prolonged illumination induced irreversible structural changes, most probably of thermo-optic origin.

Biosensing Techniques↗
PubMed2005

Porous silicon-based optical microsensor for the detection of L-glutamine.

The molecular binding between the glutamine-binding protein (GlnBP) from Escherichia coli and L-glutamine (Gln) is optically transduced by means of a biosensor based on porous silicon nano-technology. The sensor operates by the measurement of the interferometric fringes in the reflectivity spectrum of a porous silicon Fabry-Perot layer. The binding event is revealed as a shift in wavelength of the fringes. Due to the hydrophobic interaction with the Si-H terminated surface of the porous silicon, the GlnBP protein, which acts as a molecular probe for Gln, penetrates and links into the pores of the porous silicon matrix. We can thus avoid any preliminary functionalization process of the porous layer surface, which is also prevented from oxidation, at least for few cycles of wet measurements. The binding of Gln to GlnBP has also been investigated at different concentration of GlnBP.

Biosensing Techniques↗
PubMed2006

Crosslinking of phenylboronic acid receptors as a means of glucose selective holographic detection.

A holographic sensor for the detection of glucose has been developed that is based on a hydrogel film containing phenylboronic acid receptors. Changes to the replay wavelength of the hologram were used to characterise the swelling and de-swelling behaviour of the hydrogel matrix upon receptor-ligand binding. The effect of introducing a fixed positive charge into the polymer matrix by modification of the hydrogel with a quaternary amine group (3-acrylamidopropyl)trimethylammonium chloride (ATMA), was investigated for a range of sugars and the alpha-hydroxy acid, lactate, at physiological pH. The quaternary amine-modified hydrogel matrix was found to contract in the presence of glucose, whereas, it was minimally responsive to other saccharides. The selectivity of the sensor for glucose compared to lactate was also significantly improved compared to the unmodified film. A crosslinking mechanism is proposed to explain the enhanced selectivity to glucose.

Biosensing Techniques↗
PubMed2006

Registration of T-2 mycotoxin with total internal reflection ellipsometry and QCM impedance methods.

A sensitive optical method of total internal reflection ellipsometry (TIRE) in conjunction with immune assay approach was exploited for the registration of T-2 mycotoxin in a wide range of concentrations from 100 microg/ml down to 0.15 ng/ml. Association constants of 1.4x10(6) and 1.9x10(7)mol(-1)s for poly- and monoclonal T-2 antibodies, respectively, were evaluated from TIRE kinetic measurements. According to TIRE data fitting, binding of T-2 molecules to antibodies (at saturation) has resulted in the increase in adsorbed layer thickness of 4-5 nm. The QCM impedance measurements data showed anomalously large mass increase and film softening, most likely, due to the binding of large T-2 aggregates to antibodies.

Biosensing Techniques↗
PubMed2006

A reagentless optical biosensor based on the intrinsic absorption properties of peroxidase.

During the reversible reaction between peroxidase (HRP) and H(2)O(2), several peroxidase intermediate species, showing different molecular absorption spectra, are formed which can be used for H(2)O(2) determination; when H(2)O(2) is generated in a previous enzymatic reaction, the substrate involved in this reaction can also be determined. On this basis, a new family of fully reversible reagentless optical biosensors containing HRP is presented; glucose determination is used as a model. The biosensor (which can be used for at least 6 months and/or more than 750 measurements) is prepared by HRP and glucose oxidase entrapment in a polyacrylamide gel matrix. A mathematical model (in which optical, kinetic and transport aspects are considered) relating the measured absorbance with the substrate concentration is also presented together with a simple methodology for characterization of this kind of biosensor. Regarding the optical model, the Kubelka-Mulk theory of reflectance does not give good results and the biosensors are better described by the Rayleigh theory of polymer solutions. Under working conditions, linear response ranges from 1.5x10(-6) to 3.0x10(-4)M glucose and CV was about 4%. This biosensor has been applied for glucose determination in fruit juices and synthetic serum samples without sample pretreatment.

Absorption↗
PubMed2006

Deep-probe metal-clad waveguide biosensors.

Two types of metal-clad waveguide biosensors, so-called dip-type and peak-type, are analyzed and tested. Their performances are benchmarked against the well-known surface-plasmon resonance biosensor, showing improved probe characteristics for adlayer thicknesses above 150-200 nm. The dip-type metal-clad waveguide sensor is shown to be the best all-round alternative to the surface-plasmon resonance biosensor. Both metal-clad waveguides are tested experimentally for cell detection, showing a detection limit of 8-9 cells/mm2.

Biosensing Techniques↗
PubMed2006

Coupled waveguide-surface plasmon resonance biosensor with subwavelength grating.

This study develops a coupled waveguide-surface plasmon resonance (CWSPR) biosensor with a subwavelength grating structure for the real-time analysis of biomolecular interactions. In the proposed optical metrology system, normally incident white light is coupled into the waveguide layer through the subwavelength grating structure thereby enhancing the wave vector which excites the surface plasmons on the metal sensing surface. The proposed CWSPR biosensor not only retains the same sensing sensitivity as that of a conventional surface plasmon resonance device, but also yields a sharper dip in the reflectivity spectrum and therefore provides an improved measurement precision. Moreover, the metrology setup overcomes the limitations of the conventional Kretschmann attenuated total reflection approach and is less sensitive to slight variations in the angle of the incident light. The experimental results confirm that the current CWSPR biosensor provides a straightforward yet powerful technique for real-time biomolecular interaction analysis.

Biopolymers↗
PubMed2004

Studies on intestinal permeability of cirrhotic patients by analysis lactulose and mannitol in urine with HPLC/RID/MS.

The method for separation and determination of lactulose (L) and mannitol (M) in urine was developed by HPLC with a refractive index detector (RID). The linearity ranged from 5 to 1000 microg/mL for L and M, respectively. Recoveries ranged from 93.1% to 97.1%. The intra- and inter-day relative standard deviations of peak area were between 0.8-1.4% (n=3) and 1.4-3.6% (n=3). The limits of detection were obtained with 1.40 microg/mL for L and 1.65 microg/mL for M. The ratios of L/M in the urine samples for the spontaneous ascitic fluid infection (SAI), sterile ascitic fluid (SA) patients, and healthy volunteers (HV) were determined. The results showed well the correlations among the L/M ratio, intestinal permeability (IP) and the illness status of patients, and also indicated lactulose could improve the IP of SAI patients. The peaks of L and M in chromatograms were identified by electrospray ionization/mass spectrometry (ESI/MS), which ensured the accurate measurement of the ratio L/M. This method presented a rapid, accurate, and practical technique for determining IP in clinical practice and investigating the pathology of hepatocirrhosis.

Chromatography, High Pressure Liquid↗
PubMed2005

Unperturbed dimensions of Carrageenans in different salt solutions.

Commercial samples of kappa-, iota-, and lambda-carrageenans were studied by means of Size Exclusion Chromatography with dual detection, i.e. employing a Refractive Index (concentration sensitive) and Multiangle Light Scattering (size sensitive) detectors. The eluent was water containing 0.1 M concentration of different ionic salts, namely LiCl, NaCl, KCl and NaI, with the exception of kappa-carrageenan that aggregates in presence of KCl. Molecular weight distributions and averages, coefficients of the scaling law of molecular dimensions and unperturbed dimensions were thus obtained from a single polydisperse sample of each polymer. Measurements were performed at 25 degrees C and all the systems were above theta conditions with values of the q exponent of the scaling law ranging from 0.51 to 0.59. Extrapolation to unperturbed conditions provides values of the characteristic ratio C(N)=56+/-1 and 40+/-5 respectively for lambda- and iota-carrageenans regardless of the ionic salt employed. However, kappa-carrageenan gives C(N)=31, 35 and 59, respectively, in presence of LiCl, NaCl and NaI, which clearly indicates that this polymer behaves on a different way in presence of NaI than with the other two salts. A tentative explanation of this difference is provided.

Carbohydrate Conformation↗
PubMed2004

Bioactive polysaccharides from the stems of the Thai medicinal plant Acanthus ebracteatus: their chemical and physical features.

Crude water-soluble polysaccharides were isolated from Acanthus ebracteatus by hot water extraction followed by ethanol precipitation after pre-treatment with 80% ethanol. The crude polysaccharides were separated into neutral and acidic polysaccharides by anion-exchange chromatography. The neutral polysaccharide (A1001) was rich in galactose, 3-O-methylgalactose and arabinose, whereas the acidic polysaccharide (A1002) consisted mainly of galacturonic acid along with rhamnose, arabinose and galactose as minor components indicating a pectin-type polysaccharide with rhamnogalacturonan type I (RG-1) backbone. 3-O-Methylgalactose is also present in the acidic fraction. Both neutral and acidic fractions showed potent effects on the complement system using pectic polysaccharide PM II from Plantago major as a positive control. A small amount of 3-O-methylgalactose present in the pectin seemed to be of importance for activity enhancement in addition to the amount of neutral sugar side chains attached to RG-1. The relationship between chemical structure and effect on the complement system of the isolated polysaccharides is considered in the light of these data. The presence of the rare monosaccharide 3-O-methylgalactose may indicate that this can be used as a chemotaxonomic marker. The traditional way of using this plant as a medical remedy appears to have a scientific basis.

Acanthaceae↗
PubMed2004

Analysis of mono- and disaccharides in milk-based formulae by high-performance liquid chromatography with refractive index detection.

A simple and reproducible method for the qualitative and quantitative analysis of free mono- and disaccharides (fructose, glucose, galactose, sucrose, lactulose and lactose) in milk-based formulae by high-performance liquid chromatography (HPLC) with refractive index (RI) detection was developed and validated. The method showed good linearity with determination coefficients exceeding 0.99. The limits of detection (DL) in these sugars were 0.17, 0.13, 0.06, 0.16, 0.05 and 0.25 mg/ml, respectively; and the limits of quantification (QL), 0.27, 0.24, 0.20, 0.26, 0.22 and 0.38 mg/ml. The relative standard deviations (R.S.D.s) for repeatability in fructose, sucrose, lactulose and lactose were 0.78, 0.99, 2.91 and 0.46 and the R.S.D.s for reproducibility were 4.8, 6.15, 7.04 and 2.49, respectively. Recoveries in all sugars were between 93 and 113%.

Animals↗
PubMed2004

Separation of polyethylene glycols and their amino-substituted derivatives by high-performance gel filtration chromatography at low ionic strength with refractive index detection.

The chromatographic analysis of polyethylene glycols (PEG) and their amino-substituted derivatives, with an average molecular mass of 2000 and 3350 is described. The PEG derivatives were perfectly separated by TSK-GEL G4000PW(XL) column, which was widely used in high-performance size-exclusion chromatography (SEC), at low ionic strength with refractive index detection. It was shown that ion-exchange interactions were mainly involved in the retention mechanism of these compounds on TSK-GEL G4000PW(XL) column, since retention volume decreased when salt concentration were added to the mobile phase and varied with the pH of the eluent. Additionally, size exclusion for PEG chains plays a role. Organic modifiers also had effect on chromatographic behaviors of PEG compounds.

Amines↗
PubMed2005

Column selectivity in reversed-phase liquid chromatography. VIII. Phenylalkyl and fluoro-substituted columns.

As reported previously, five solute-column interactions (hydrophobicity, steric resistance, hydrogen-bond acidity and basicity, ionic interaction) quantitatively describe column selectivity for 163 alkyl-silica, polar-group and cyano columns. In the present study, solute retention and column selectivity for 11 phenyl and 5 fluoro-substituted columns were compared with alkyl-silica columns of similar ligand length. It is concluded that two additional solute-column interactions may be significant in affecting retention and selectivity for the latter columns: (a) dispersion interactions of varying strength as a result of significant differences in bonded-phase polarizability or refractive index and (b) pi-pi interactions in the case of phenyl columns and aromatic solutes. These 16 phenyl and fluoro columns were also characterized in terms of hydrophobicity, steric resistance, hydrogen-bond acidity and basicity, and ionic interaction.

Chromatography, Liquid↗
PubMed2005

Distribution analysis of ultra-high molecular mass poly(ethylene oxide) containing silica particles by size-exclusion chromatography with dual light-scattering and refractometric detection.

Two different size-exclusion chromatography (SEC) systems, connected in-line either to a low-angle light scattering (LALS) or to a multiangle light scattering (MALS) detector, are employed for determination of molecular mass distributions (MMD) of poly(ethylene oxide) (PEO) samples having a weight average molecular mass up to eight millions. The detrimental effect of the presence of strongly scattering silica particles in the samples on the light scattering signal can be eliminated using a suitable sample dissolution procedure utilizing silica solubility in aqueous mobile phase. The selection of flow-rate and sample concentration have a large impact on the obtained results. Hydrodynamic retardation phenomena and nonlinearity effects are shown to introduce severe errors in the molecular mass distributions unless flow-rate and sample concentration are kept at sufficiently low levels. Self-compensating ability of the dual detection in flow-rate effects is shown to be the main advantage here. A good agreement between the results obtained using LALS and MALS detection is found provided that a carefully selected angular extrapolation procedure is used in the case of MALS data. Thus, using carefully selected experimental conditions, SEC with light-scattering (LS) and refractometric detection proved to be an efficient technique for MMD characterisation also of ultra-high molecular mass (UHM) PEO polymers.

Chromatography, Gel↗