Showing papers by "David E. Cliffel published in 2005"

Quartz crystal microbalance detection of glutathione-protected nanoclusters using antibody recognition.

Abstract: A quartz crystal microbalance (QCM) immunosensor was developed for the quantitative detection of glutathione-protected nanoclusters. Advantages intrinsic to QCM were employed to make it an attractive alternative to other immunosensing techniques. We have addressed challenges in the area of QCM mass sensing through experimental correlation between damping resistance and frequency change for a reliable mass measurement. Electrode functionalization was optimized with the use of protein A to immobilize and present polyclonal IgG for antigen binding. This method was developed for the detection of glutathione (antigen)-protected clusters of nanometer size with high surface area and thiolate valency. Quantitation of glutathione−nanocluster binding to immobilized polyclonal antibody provides equilibrium constants (Ka = (3.6 ± 0.2) × 105 M-1) and kinetic rate constants (kf = (5.4 ± 0.7) × 101 M-1 s-1 and kr = (1.5 ± 0.4) × 10-4 s-1) comparable to literature reports. These observations further imply that immunoreac...

Photosystem I patterning imaged by scanning electrochemical microscopy.

Abstract: We report the first directed adsorption of Photosystem I (PSI) on patterned surfaces containing discrete regions of methyl- and hydroxyl-terminated self-assembled monolayers (SAMs) on gold. SAM and PSI patterns are characterized by scanning electrochemical microscopy (SECM). The insulating protein complex layer blocks the electron transfer of the SECM mediator, thereby reducing the electrochemical current significantly. Uniformly and densely packed adsorbed protein layers are observed with SECM. Pattern images correlate with our previous studies where we showed that low-energy surfaces (e.g., CH3-terminated) inhibit PSI adsorption in the presence of Triton X-100, whereas high-energy surfaces (e.g., OH-terminated) enable adsorption. Therefore, a SAM pattern with alternating methyl and hydroxyl surface regions allows PSI adsorption only on the hydroxyl surface, and this is demonstrated in the resulting SECM images.

Continuous free-flow electrophoresis of water-soluble monolayer-protected clusters.

Abstract: There has recently been a surge of interest in the properties and applications of monolayer protected clusters (MPCs). MPCs are metal nanoparticles that have unique optical, chemical, and electrochemical properties resulting from their small size. Because the size defines their properties, MPC particle size fractionation is important for control of the MPC characteristics for use in many potential applications. This paper explores the use of continuous free-flow electrophoresis (CFE) for the size fractionation of N-(2-mercaptopropionyl)glycine (tiopronin) monolayer protected gold clusters into monodisperse nanoparticle samples. CFE is a fractionation technique that isolates monodisperse particle sizes into several different collection vials on the tens of milligrams scale. This allows the MPCs to be separated based on their electrophoretic mobilities into isolated, monodisperse particles across a wide range of sizes. CFE separation of water-soluble tiopronin MPCs yielded fractions that varied in color, UV-visible spectra, transmission electron microscopy (TEM) size histograms, and solubility, indicating narrow size dispersity in the isolated fractions. UV-visible spectrophotometry verified the separation of the tiopronin MPCs through the inspection of surface plasmon resonance peak sizes for the different fractions. TEM was also used to verify the narrowed dispersity of MPC samples. The ability to separate water-soluble nanoparticles into 30 or more fractions in a continuous flow process will enable future studies on their size dependent properties.

Real-time cell dynamics with a multianalyte physiometer.

Abstract: A technique for simultaneously measuring changes in extracellular glucose, lactate, and oxygen concentrations in conjunction with acidification rates on a Cytosensor Microphysiometer is described. Platinum electrodes are inserted into the standard Cytosensor plunger head and modified with enzymes and biocompatible polymeric films. The lactate and glucose oxidase enzymes catalyze the reaction of lactate and glucose. An end product of these catalyses, H2O2, is measured amperometrically. Extracellular oxygen is also measured amperometrically, while the acidification rate is measured potentiometrically by the Cytosensor. Useful information is obtained during the Cytosensor stop-flow cycles, which produce increasing or decreasing peaks, owing to the production of lactic and carbonic acid and consumption of glucose and oxygen by the cells. Fabrication of the modified sensor head and deposition of the electrode films is detailed, and the operation of the technique is described and illustrated by the simultaneous measurement of all four analytes during the addition of 20 mM fluoride to mouse fibro blast cells.