Glycosphingolipids (GSLs) have been shown to undergo strong interactions with a number of protein toxins, including potential bioterrorism agents such as ricin and botulinum neurotoxin. Characterization of this interaction in recent years has led to a number of studies where GSLs were used as the recognition molecules for biosensing applications. Here, we offer a comparison of quartz crystal microbalance (QCM) sensors for the detection of ricin using antibodies and the GSLs GM1 and asialoGM1, which have been shown to undergo strong interactions with ricin. The presence, orientation, and activity of the GSL and antibody films were confirmed using ellipsometry, Fourier transform infrared spectroscopy (FT-IR), and QCM. It was found that the GSLs offered more sensitive detection limits when directly compared with antibodies. Both GSLs had lower detection limits at 5 microg/mL, approximately 5 times lower than were found for antibodies (25 microg/mL), and their linear detection range extended to the highest concentrations tested (100 microg/mL), almost an order of magnitude beyond the saturation point for the antibody sensors. Potential sites for nonspecific adsorption were blocked using serum albumin without sacrificing toxin specificity.
"Ricin is one of the most toxic and easily obtainable plant toxins and is isolated from the seeds of castor bean Ricinus communis (Roberts and Smith, 2004). The ricin toxin is a heterodimer comprised of a 32 kDa A chain and a 34 kDa B chain connected by interchain disulfide bonds (Stine et al., 2005). The B chain is a lectin which binds specifically to galactosyl residues on the cell surface and appears to trigger the endocytic uptake of ricin. "
[Show abstract][Hide abstract] ABSTRACT: We developed a novel silica coating magnetic nanoparticle-based silver enhancement immunoassay (SEIA) for ricin toxin (RT) rapid electrical detection using interdigitated array microelectrodes (IDAMs) as electrodes. This novel system was developed by taking advantage of the separation and enrichment properties of magnetic nanoparticles (MNPs) and the catalytic properties of gold nanoparticles (GNPs). In this system, MNPs labeled with anti-ricin A chain antibody 6A6 were used to capture ricin and GNPs labeled with anti-ricin B chain antibody 7G7 were used as detectors. To enhance the electrical signal, the catalytic properties of GNPs were used to promote silver reduction. In the presence of ricin, a sandwich structure was formed which could be separated by a magnetic field. The sandwich complex was then transferred to IDAMs. The silver particles bridged the IDAM gaps and gave rise to an enhancing electrical signal that was detected by conductivity measurements. The results showed that the sensitivity of the SEIA for ricin electrical detection was five times greater than that of conventional colorimetric sandwich ELISA. Once the antibody used for detection was coated on the plates or MNPs, our system was three times more rapid than colorimetric sandwich ELISA. This rapid and sensitive detection system provides promising new potential for ricin detection.
[Show abstract][Hide abstract] ABSTRACT: Development of simple and painless techniques of monitoring metabolites like glucose with increased frequency would be beneficial to diabetic patients. Implantable sensors for glucose have been under investigation for nearly three decades, Poor stability and sensitivity of these sensors limit their use in closed-loop delivery. In this work we have taken advantage of silicon micro-fabrication technologies to develop implantable redundant microsensor arrays with glucose oxidase molecules immobilized in photopolymerized and microlithographically patterned films. We have used redox polymers that exchange electrons with glucose oxidase and also form macromolecular networks with these enzymes. The enzymes entrapped in these polymer films and containing biocompatible hydrogels show good stability and sensitivity. Key accomplishments include: a)Successively synthesized an osmium based polycationic redox polymer (POs-Ea) a molecule that is responsible for exchanging electrons with glucose oxidase enzyme; b) Used photolithography to fabricate patterned sensor arrays on flexible plastic substrates (mylar and polyimide); c) Successively crosslinked active glucose oxidase enzyme with redox polymer and biocompatible polyethylene glycol diacrylate hydrogel; d)Used amperometry and cyclic voltammetry to confirm activity of the enzyme and contribution of each sensor array element; e) The enzyme exchanged electrons with redox polymer both entrapped in a hydrogel network.
[Show abstract][Hide abstract] ABSTRACT: Phospholipid films have been shown in a number of studies to exhibit potential as nonfouling surfaces for biomaterial applications. However, the practical application of such films has been hindered by instability in aqueous solutions and significant detachment under mild shear stresses. Methods for stabilizing lipid films have been investigated, but to date require the presence of specific functional groups or chemical modification of the lipid molecule. In contrast to these methods, we present a process for heat-stabilization of lipid films. These heat-stabilized films have been shown to be able to withstand repeated rinsing without significant detachment. Phosphatidylcholine monolayers were formed on hydrophobic self-assembled monolayers using the liposome fusion method and stabilized at 80 degrees C. The films were characterized using Fourier transform infrared spectroscopy, ellipsometry, and atomic force microscopy and were shown to be defect free after repeated rinsing. Further experiments using a quartz crystal microbalance showed that the heat-stabilized lipid films were highly resistant to nonspecific protein adhesion and compared very favorably with poly(ethylene glycol)-coated surfaces under identical exposure conditions.
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