FG Nanobiotechnologie
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Institute
Bimetallic layers increase sensitivity of affinity sensors based on surface plasmon resonance
(2002)
Two metals are used in resonant layers for chemical sensors based on surface plasmon resonance (SPR) - gold and silver. Gold displays higher shift of the resonance angle to changes of ambient refraction index and is chemically stable. Silver posses narrower resonance curve thus providing a higher signal/noise ratio of SPR chemical sensors, but has a poor chemical stability. A new structure of resonant metallic film based on bimetallic silver/gold layers (gold as an outer layer) is suggested. It combines advantages of both gold and silver resonant layers. Bimetallic resonant films display so high shift of resonance angle on changes of ambient refraction index as gold films, but show narrower resonance curve, thus providing a higher signal / noise ratio. Additionally, the outer gold layer protects silver against oxidation.
A new technique for on-line monitoring of analyte binding to sensor surfaces by surface plasmon resonance (SPR) detection is described. It is based on differential measurements using two wavelengths provided by two diode lasers. The technique is as simple and robust as the conventional SPR detection measuring the reflected radiation at fixed incidence angle, but it has the advantage of being nonsensitive to variations of the resonance width and providing essentially higher signal/noise ratios. The paper presents the first four channel prototype system for parallel 2D-monitoring at four different spots. One channel is always used as a reference to compensate temperature fluctuations and nonspecific adsorptions. Calibration with sucrose solutions revealed an absolute sensitivity of Δn ∼ 5 × 10-6. The new technique is tested with a biotin−streptavidin binding and with hybridization/denaturation of DNA. Biotin binding to a streptavidin monolayer is detected with a signal/noise ratio of about 5, which demonstrates the high potential of the new technique for applications in drug discovery. Applications to gene analysis are tested with short oligonucleotides of the sequences used for genotyping human hepatitis C viruses. A selective response to complementary oligonucleotides is observed. The high reproducibility in subsequent cycles of hybridization/denaturation (by formamide or by heating) points out potential applications of the technique in medical diagnostics, food industry, genomics, and proteomics too.
The detection limit of surface plasmon resonance (SPR) measurements has been improved by a factor of approximately 2-3.5 if the angle of incidence was near to the reflection minimum of the SPR resonance curve instead at the position of the steepest slope, the standard alignment in SPR imaging. The enhancement of the detection power, a result of signal-to-noise optimization, is demonstrated by applying a photodiode and a CCD camera for SPR detection. The experimental data are compared with data expected from theory.
Electron transfer between metal electrodes and ferro/ferricyanide is completely suppressed at low ionic concentration. We describe here a new phenomenon related to this reaction: an immobilisation of thiolated single-stranded DNA on gold electrodes retains this activity at low ionic strength up to the level corresponding to the high ionic strength. In contrast, a hybridisation of the complementary DNAwith the thiolated single-stranded DNA followed by a binding onto the electrodes, attenuated the electrocatalytic effect. These effects can be used for discrimination between single-stranded DNA and double-stranded DNA and for semi-quantitative measurement of complementary DNA in a sample.
In this research, a surface-plasmon resonance biosensor for nucleic acid detection was constructed, optimized and characterized. The kinetic response on hybridization was also studied. The biosensor was use as a platform for the study of the surface coverage control by single-strand or double-strand oligonucleotide immobilization. The dsDNA immobilization and offers a new way to surface coverage control for the nucleic acid biosensor construction.
Carbodiimides possess considerable absorbance in the ultraviolet region; the extinction coefficient of 1-ethyl-(3-dimethylaminopropyl)carbodiimide (EDC) in water is ϵ214 = 6.3 · 103 L mol−1 cm−1. It provides a very simple method for testing possible side reactions of carbodiimides. This technique was used to study effects of pH, different buffers, and other components typically present in biological samples on EDC stability. It was shown that the hydrolysis rate in pure water increased from 1.5 · 10−5 to 5 · 10−4 s−1 by pH decreasing from 7 to 4. A strong increase of the rate of EDC loss was observed in the presence of different components. This effect can be described by the following row of hydrolysis rates: citrate ⪢ acetate ≈ phosphate ⪢ SDS. The results can be used to optimize carbodiimide-mediated reactions of peptide bond formation in organic chemistry or peptide synthesis or during immobilization or cross-linking of biological molecules.
32P-labeled DNA oligomers were immobilized onto self-assembled monolayers formed by carboxy-modified alkylthiols on gold electrodes. The optimal conditions were evaluated by comparing of surface density of oligonucleotides measured for different immobilization conditions before and after desorption of physically adsorbed nucleotides. The results demonstrate that the physical adsorption depends strongly on immobilization conditions being typically 10 times higher than the chemical immobilization. Under optimal immobilization conditions, the surface density of oligonucleotides was more than 20% of the value calculated for a densely packed monolayer.
Changes in the electrical conductance at 240 °C of a number of copper(I) compounds including CuI, CuBr, Cu2Te, Cu12Sb4S13, (CuI)2P14, and of NiO were measured following the exposure to the gases ammonia, nitrogen dioxide, nitrous oxide, carbon monoxide, carbon dioxide, and propane. Analytical sensitivity and kinetic parameters were compared quantitatively. The high diversity of sensitivity patterns makes these compounds promising candidates for use in sensor arrays. Principal component analysis of the data obtained with a virtual sensor array enabled highly selective sensing of five of the six analytes studied using only two principle components. Activation energies of the charge transport in the materials were determined.