Analysis of Protein Adsorption and Binding at Biosensor Polymer Interfaces Using X-ray Photon Spectroscopy and Scanning Electrochemical Microscopy

Abstract

We describe a method, based on X-ray photoelectron spectroscopy (XPS) measurements, to assess the extent of protein adsorption or binding on a variety of different muTAS and biosensor interfaces. Underpinning this method is the labeling of protein molecules with either iodine- or bromine-containing motifs by using protocols previously developed for radiotracer studies. Using this method, we have examined the adsorption and binding properties of a variety of modified electrodeposited polymer interfaces as well as other materials used in muTAS device fabrication. Using polymer interfaces modified with poly(propylene glycol) (PPG) chains, our results indicate that a chain of at least approximately 30 monomer units is required to inhibit nonspecific adsorption from concentrated protein solutions. The XPS methodology was also used to probe specific binding of avidins and enzyme conjugates thereof to biotinylated and mixed biotin/PPG-modified polymer interfaces. In one example, using competitive binding, it was established that the mode of binding of a peroxidase-streptavidin conjugate to a biotinylated modified polymer interface was primarily via the streptavidin moiety (as opposed to nonspecific binding via the enzyme conjugate). XPS evaluation of nonspecific and specific peroxidase-streptavidin immobilization on various functionalized polymers has guided the design and fabrication of functionalized interdigitated electrodes in a biosensing muTAS device. Subsequent characterization of this device using scanning electrochemical microscopy (SECM) corroborated the adsorption and binding previously inferred from XPS measurements on macroscale electrodes.

Influence of Iodide Ions on the Growth of Gold Nanorods: Tuning Tip Curvature and Surface Plasmon Resonance

“…6B), the I-3d 5/2 peak also shows two contributions at 620.1 and 619.4 eV, which can be unambiguously assigned to gold and silver iodide, respectively. [24,25] Notably, the silver halide signal is in both cases much larger than that of gold, suggesting that Ag ions are concentrated at the surface.…”

Noncovalent Immobilization of Streptavidin on In Vitro- and In Vivo-Biotinylated Bacterial Magnetic Particles

“…Although various methods for immobilizing streptavidin on solid surfaces have been reported, the streptavidin-biotin interaction is the most commonly used method (8,13,32). In this method, the solid surface is first biotinylated using a crosslinker reagent, and this is followed by the addition of streptavidin.…”

The immobilization of proteins on biodegradable polymer fibers via click chemistry

“…These results indicate the effectiveness of blocking, on one hand, and demonstrate the necessity of blocking on each layer of immobilized or bound proteins, on other hand. There are other possibilities for non-specific adsorptions [35]. For example, in the reaction of immobilized TSP50 with anti-TSP50, if the immobilized TSP50 did not entirely react with anti-TSP50, it would be left free on the surface and cause subsequent undesirable adsorption of HRP-IgG.…”