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Review of Bioassays for Monitoring Fate and Transport ofEstrogenic Endocrine Disrupting Compounds in Water

Description: Endocrine disrupting compounds (EDCs) are recognizedcontaminants threatening water quality. Despite efforts in sourceidentification, few strategies exist for characterization or treatment ofthis environmental pollution. Given that there are numerous EDCs that cannegatively affect humans and wildlife, general screening techniques likebioassays and biosensors provide an essential rapid and intensiveanalysis capacity. Commonly applied bioassays include the ELISA and YESassays, but promising technologies include ER-CALUXa, ELRA, Endotecta,RIANA, and IR-bioamplification. Two biosensors, Endotecta and RIANA, arefield portable using non-cellular biological detection strategies.Environmental management of EDCs in water requires integration ofbiosensors and bioassays for monitoring and assessment.
Date: January 30, 2004
Creator: CGCampbell@lbl.gov
Partner: UNT Libraries Government Documents Department

Field-Portable Immunoassay Instruments and Reagents to Measure Chelators and Mobile Forms of Uranium

Description: Previous studies from our laboratory have demonstrated the feasibility of immunoassays for identification and quantification of specific metal ions. Our ultimate goal for this project is to (1) isolate and characterize antibodies that recognize the most mobile form of uranium, UO22+; (2) assemble, test, and validate a new field-portable immunosensor based on these antibodies; (3) prepare new monoclonal antibodies to the primary chelators (EDTA and DTPA) found in DOE wastes.
Date: June 1, 2001
Creator: Blake, Diane A.
Partner: UNT Libraries Government Documents Department

Field-based detection and monitoring of uranium in contaminated groundwater using two immunosensors

Description: Field-based monitoring of environmental contaminants has long been a need for environmental scientists. Described herein are two kinetic exclusion-based immunosensors, a field portable sensor (FPS) and an inline senor, that were deployed at the Integrated Field Research Challenge Site of the U.S. Department of Energy in Rifle, CO. Both sensors utilized a monoclonal antibody that binds to a U(VI)-dicarboxyphenanthroline complex (DCP) in a kinetic exclusion immunoassay format. These sensors were able to monitor changes of uranium in groundwater samples from {approx} 1 {micro}M to below the regulated drinking water limit of 126 nM (30 ppb). The FPS is a battery-operated sensor platform that can determine the uranium level in a single sample in 5-10 min, if the instrument has been previously calibrated with standards. The average minimum detection level (MDL) in this assay was 0.33 nM (79 ppt), and the MDL in the sample (based on a 1:200?1:400 dilution) was 66?132 nM (15.7?31.4 ppb). The inline sensor, while requiring a grounded power source, has the ability to autonomously analyze multiple samples in a single experiment. The average MDL in this assay was 0.12 nM (29 ppt), and the MDL in the samples (based on 1:200 or 1:400 dilutions) was 24?48 nM (5.7?11.4 ppb). Both sensor platforms showed an acceptable level of agreement (r{sup 2} = 0.94 and 0.76, for the inline and FPS, respectively) with conventional methods for uranium quantification.
Date: May 1, 2009
Creator: Melton, S.J.; Yu, H.; Williams, K.H.; Morris, S.A.; Long, P.E. & Blake, D.A.
Partner: UNT Libraries Government Documents Department

Polycyclic aromatic hydrocarbons at selected burning grounds at Los Alamos National Laboratory

Description: A commercial immunoassay field test (IFT) was used to rapidly assess the total concentrations of polycyclic aromatic hydrocarbons (PAHs) in the soil at selected burning grounds within the explosives corridor at Los Alamos National Laboratory (LANL). Results were compared with analyses obtained from LANL Analytical Laboratory and from a commercial laboratory. Both used the Environmental Protection Agency`s (EPA`s) Methods 8270 and 8310. EPA`s Method 8270 employs gas chromatography and mass spectral analyses, whereas EPA`s Method 8310 uses an ultraviolet detector in a high-performance liquid chromatography procedure. One crude oil sample and one diesel fuel sample, analyzed by EPA Method 8270, were included for references. On an average the IFT results were lower for standard samples and lower than the analytical laboratory results for the unknown samples. Sites were selected to determine whether the PAHs came from the material burned or the fuel used to ignite the burn, or whether they are produced by a high-temperature chemical reaction during the burn. Even though the crude oil and diesel fuel samples did contain measurable quantities of PAHs, there were no significant concentrations of PAHs detected in the ashes and soil at the burning grounds. Tests were made on fresh soil and ashes collected after a large burn and on aged soil and ashes known to have been at the site more than three years. Also analyzed were twelve-year-old samples from an inactive open burn cage.
Date: February 1, 1998
Creator: Harris, B.W.; Minor, L.K.M. & Flucas, B.J.
Partner: UNT Libraries Government Documents Department

LDRD final report on microencapsulated immunoreagents for development of one-step ELISA

Description: Microencapsulation of biological macromolecules was investigated as a method for incorporating the necessary immunoreagents into an improved enzyme-linked immunosorbant assay (ELISA) package that would self-develop. This self-contained ELISA package would eliminate the need for a trained technician to perform multiple additions of immunoreagent to the assay. Microencapsulation by insolution drying was selected from the many available microencapsulation methods, and two satisfactory procedures for microencapsulation of proteins were established. The stability and potential for rapid release of protein from these microencapsulates was then evaluated. The results suggest that the chosen method for protein entrapment produces microcapsules with a considerable amount of protein in the walls making these particular microcapsules unsuitable for their intended use.
Date: August 1, 1997
Creator: Henderson, C.C. & Singh, A.K.
Partner: UNT Libraries Government Documents Department

Towards Chip Scale Liquid Chromatography and High Throughput Immunosensing

Description: This work describes several research projects aimed towards developing new instruments and novel methods for high throughput chemical and biological analysis. Approaches are taken in two directions. The first direction takes advantage of well-established semiconductor fabrication techniques and applies them to miniaturize instruments that are workhorses in analytical laboratories. Specifically, the first part of this work focused on the development of micropumps and microvalves for controlled fluid delivery. The mechanism of these micropumps and microvalves relies on the electrochemically-induced surface tension change at a mercury/electrolyte interface. A miniaturized flow injection analysis device was integrated and flow injection analyses were demonstrated. In the second part of this work, microfluidic chips were also designed, fabricated, and tested. Separations of two fluorescent dyes were demonstrated in microfabricated channels, based on an open-tubular liquid chromatography (OT LC) or an electrochemically-modulated liquid chromatography (EMLC) format. A reduction in instrument size can potentially increase analysis speed, and allow exceedingly small amounts of sample to be analyzed under diverse separation conditions. The second direction explores the surface enhanced Raman spectroscopy (SERS) as a signal transduction method for immunoassay analysis. It takes advantage of the improved detection sensitivity as a result of surface enhancement on colloidal gold, the narrow width of Raman band, and the stability of Raman scattering signals to distinguish several different species simultaneously without exploiting spatially-separated addresses on a biochip. By labeling gold nanoparticles with different Raman reporters in conjunction with different detection antibodies, a simultaneous detection of a dual-analyte immunoassay was demonstrated. Using this scheme for quantitative analysis was also studied and preliminary dose-response curves from an immunoassay of a mo del antigen were obtained. Simultaneous detection of several analytes at the same address can potentially increase the analysis speed, and can further expand the analysis capability of a microarray chip.
Date: September 21, 2000
Creator: Ni, J.
Partner: UNT Libraries Government Documents Department

Evaluation of ELISA screening test for detecting aflatoxin in biogenic dust samples

Description: Aflatoxin is a carcinogenic chemical that is sometimes produced when agricultural commodities are infested by the fungi Aspergillus flavus and A. Parasiticus. Aflatoxin has been found to be present in air samples taken around persons handling materials likely to be contaminated. The purpose of this investigation was to demonstrate the feasibility of using an Enzyme Linked Immunosorbent Assay (ELISA) test kit that was developed to screen for aflatoxin in bulk agricultural commodities, to an air sample. Samples were taken from two environments likely to be contaminated with aflatoxin, a dairy farm feed mixing operation and a peanut bagging operation. The dust collected from these environments was considered to be biogenic, in that it originated primarily from biological materials.
Date: May 1, 1996
Creator: Durant, J.T.
Partner: UNT Libraries Government Documents Department

Silver enhancement of nanogold and undecagold

Description: A recent advance in immunogold technology has been the use of molecular gold instead of colloidal gold. A number of advantages are realized by this approach, such as stable covalent, site-specific attachment, small probe size and absence of aggregates for improved penetration. Silver enhancement has led to improved and unique results for electron and light microscopy, as well as their use with blots and gels. Most previous work with immunogold silver staining has been done with colloidal gold particles. More recently, large gold compounds (``clusters``) having a definite number of gold atoms and defined organic shell, have been used, frequently with improved results. These gold dusters, large compared to simple compounds, are, however, at the small end of the colloidal gold scale in size; undecagold is 0.8 nm and Nanogold is 1.4 nm. They may be used in practically all applications where colloidal gold is used (Light and electron microscopy, dot blots, etc.) and in some unique applications, where at least the larger colloidal golds don`t work, such as running gold labeled proteins on gels (which are later detected by silver enhancement). The main differences between gold clusters and colloidal golds are the small size of the dusters and their covalent attachment to antibodies or other molecules.
Date: July 1, 1995
Creator: Hainfield, J.F. & Furuya, F.R.
Partner: UNT Libraries Government Documents Department

Evaluation of Multiplexed Foot-and-Mouth Disease Nonstructural Protein Antibody Assay Against Standardized Bovine Serum Panel

Description: Liquid array technology has previously been used to show proof-of-principle of a multiplexed non structural protein serological assay to differentiate foot-and-mouth infected and vaccinated animals. The current multiplexed assay consists of synthetically produced peptide signatures 3A, 3B and 3D and recombinant protein signature 3ABC in combination with four controls. To determine diagnostic specificity of each signature in the multiplex, the assay was evaluated against a naive population (n = 104) and a vaccinated population (n = 94). Subsequently, the multiplexed assay was assessed using a panel of bovine sera generated by the World Reference Laboratory for foot-and-mouth disease in Pirbright, UK. This sera panel has been used to assess the performance of other singleplex ELISA-based non-structural protein antibody assays. The 3ABC signature in the multiplexed assay showed comparative performance to a commercially available non-structural protein 3ABC ELISA (Cedi test{reg_sign}) and additional information pertaining to the relative diagnostic sensitivity of each signature in the multiplex is acquired in one experiment. The encouraging results of the evaluation of the multiplexed assay against a panel of diagnostically relevant samples promotes further assay development and optimization to generate an assay for routine use in foot-and-mouth disease surveillance.
Date: May 14, 2007
Creator: Perkins, J; Parida, S & Clavijo, A
Partner: UNT Libraries Government Documents Department

Grande Ronde Basin Spring Chinook Salmon Captive Broodstock Program, 2008 Annual Report.

Description: The Grande Ronde Basin Spring Chinook Salmon Captive Broodstock Program is designed to rapidly increase numbers of Chinook salmon in stocks that are in imminent danger of extirpation in Catherine Creek (CC), Lostine River (LR) and upper Grande Ronde River (GR). Natural parr are captured and reared to adulthood in captivity, spawned (within stocks) and their progeny reared to smoltification before being released into the natal stream of their parents. This program is co-managed by ODFW, National Marine Fisheries Service, Nez Perce Tribe and Confederated Tribes of the Umatilla Indian Reservation. Presmolt rearing was initially conducted at Lookingglass Fish Hatchery (LFH) but parr collected in 2003 and later were reared at Wallowa Fish Hatchery (WFH). Post-smolt rearing is conducted at Bonneville Fish Hatchery (BOH - freshwater) and at Manchester Research Station (MRS - saltwater). The CC and LR programs are being terminated, as these populations have achieved the goal of a consistent return of 150 naturally spawning adults, so the 2005 brood year was the last brood year collected for theses populations. The Grande Ronde River program continued with 300 fish collected each year. Currently, we are attempting to collect 150 natural parr and incorporate 150 parr collected as eggs from females with low ELISA levels from the upper Grande Ronde River Conventional Hatchery Program. This is part of a comparison of two methods of obtaining fish for a captive broodstock program: natural fish vs. those spawned in captivity. In August 2007, we collected 152 parr (BY 2006) from the upper Grande Ronde River and also have 155 Grande Ronde River parr (BY 2006) that were hatched from eyed eggs at LFH. During 2008, we were unable to collect natural parr from the upper Grande Ronde River. Therefore, we obtained 300 fish from low ELISA females from the upper Grande ...
Date: March 31, 2009
Creator: Hoffnagle, Timothy L.; Hair, Donald & Gee, Sally
Partner: UNT Libraries Government Documents Department

Field-Portable Immunoassay Instruments and Reagents to Measure Chelators and Mobile Forms of Uranium

Description: A collaborator in the Chemistry Department at Tulane University, Dr. Harry Ensley, has synthesized a new bifunctional chelator with specificity for ionic mercury (Hg2+) and methymercury (MeHg+). These chelators are based upon phenantholine derivative containing sulfhydryl functional groups. Experiments are underway to generate protein conjugates of this chelator to use in immunizations and screening.
Date: June 1, 2005
Creator: Blake, Diane A.
Partner: UNT Libraries Government Documents Department

Direct immunofluorescence and enzyme-linked immunosorbent assays for evaluating chlorinated hydrocarbon degrading bacteria

Description: Immunological procedures were developed to enumerate chlorinated hydrocarbon degrading bacteria. Polyclonal antibodies (Pabs) were produced by immunizing New Zealand white rabbits against 18 contaminant-degrading bacteria. These included methanotrophic and chlorobenzene (CB) degrading species. An enzyme-linked immunosorbent assay (ELISA) was used to test for specificity and sensitivity of the Pabs. Direct fluorescent antibodies (DFAs) were developed with these Pabs against select methanotrophic bacteria isolated from a trichloroethylene (TCE) contaminated landfill at the Savannah River Site (SRS) and cultures from the American Type Culture Collection (ATCC). Analysis of cross reactivity testing data showed some of the Pabs to be group specific while others were species specific. The threshold of sensitivity for the ELISA is 105 bacteria cells/ml. The DFA can detect as few as one bacterium per ml after concentration. Results from the DFA and ELISA techniques for enumeration of methanotrophic bacteria in groundwater were higher but not significantly different (P < 0.05) compared to indirect microbiological techniques such as MPN. These methods provide useful information on in situ community structure and function for bioremediation applications within 1--4 hours of sampling.
Date: June 1, 1997
Creator: Brigmon, R.L.; Franck, M.M.; Brey, J.; Fliermans, C.B.; Scott, D. & Lanclos, K.
Partner: UNT Libraries Government Documents Department

Application of fluorescent antibody and enzyme-linked immunosorbent assays for TCE and PAH degrading bacteria

Description: Historically, methods used to identify methanotrophic and polyaromatic hydrocarbon-degrading (PAH) bacteria in environmental samples have been inadequate because isolation and identification procedures are time-consuming and often fail to separate specific bacteria from other environmental microorganisms. Methanotrophic bacteria have been isolated and characterized from TCE-contaminated soils (Bowman et al. 1993; Fliermans et al., 1988). Fliermans et al., (1988) and others demonstrated that cultures enriched with methane and propane could cometabolically degrade a wide variety of chlorinated aliphatic hydrocarbons including ethylene; 1,2-cisdichloroethylene (c-DCE); 1,2-trans-dichloroethylene (t-DCE); vinyl chloride (VC); toluene; phenol and cresol. Characterization of select microorganisms in the natural setting is important for the evaluation of bioremediation potential and its effectiveness. This realization has necessitated techniques that are selective, sensitive and easily applicable to soils, sediments, and groundwater (Fliermans, et al., 1994). Additionally these techniques can identify and quantify microbial types in situ in real time
Date: July 1, 1996
Creator: Brigmon, R.L.; Franck, M.; Brey, J.; Scott, D.; Lanclos, K. & Fliermans, C.
Partner: UNT Libraries Government Documents Department

Giant Magnetoresistive Sensors and Magnetic Labels for Chip-Scale Detection of Immunosorbent Assays

Description: The combination of giant magnetoresistive sensors, magnetic labeling strategies, and biomolecule detection is just beginning to be explored. New readout methods and assay formats are necessary for biomolecules detection to flourish. The work presented in this dissertation describes steps toward the creation of a novel detection method for bioassays utilizing giant magnetoresistive sensors as the readout method. The introduction section contains a brief review of some of the current methods of bioassay readout. The theoretical underpinnings of the giant magnetoresistive effect are also discussed. Finally, the more prominent types of giant magnetoresistive sensors are described, as well as their complicated fabrication. Four data chapters follow the introduction; each chapter is presented as a separate manuscript, either already published or soon to be submitted. Chapter 1 presents research efforts toward the production of a bioassay on the surface of a gold-modified GMR sensor. The testing of this methodology involved the capture of goat a-mouse-coated magnetic nanoparticles on the mouse IgG-modified gold surface. The second, third and fourth chapters describe the utilization of a self-referenced sample stick for scanning across the GMR sensor. The sample stick consisted of alternating magnetic reference and bioactive gold addresses. Chapter 2 is concerned with the characterization of both the scanning readout method and the binding and detection of streptavidin-coated magnetic particles to a biotinylated surface. Chapter 3 advances the sample stick readout with the use of the system for detection of a sandwich immunoassay with rabbit IgG proteins. Finally, simultaneous detection of three IgG proteins is demonstrated in Chapter 4. The dissertation is concluded with a brief summary of the research presented and a discussion of the possible future applications and direction of this work.
Date: December 17, 2005
Creator: Millen, Rachel Lora
Partner: UNT Libraries Government Documents Department

Laboratory Tests of Multiplex Detection of PCR Amplicons Using the Luminex 100 Flow Analyzer

Description: Lawrence Livermore National Laboratory (LLNL) demonstrated the power of flow cytometry in detecting the biological agents simulants at JFT III. LLNL pioneered in the development of advanced nucleic acid analyzer (ANM) for portable real time identification. Recent advances in flow cytometry provide a means for multiplexed nucleic acid detection and immunoassay of pathogenic microorganisms. We are presently developing multiplexed immunoassays for the simultaneous detection of different simulants. Our goal is to build an integrated instrument for both nucleic acid analysis and immuno detection. In this study we evaluated the Luminex LX 100 for concurrent identification of more than one PCR amplified product. ANAA has real-time Taqman fluorescent detection capability for rapid identification of field samples. However, its multiplexing ability is limited by the combination of available fluorescent labels. Hence integration of ANAA with flow cytometry can give the rapidity of ANAA amplification and the multiplex capability of flow cytometry. Multiplexed flow cytometric analysis is made possible using a set of fluorescent latex microsphere that are individually identified by their red and infrared fluorescence. A green fluorochrome is used as the assay signal. Methods were developed for the identification of specific nucleic acid sequences from Bacillus globigii (Bg), Bacillus thuringensis (Bt) and Erwinia herbicola (Eh). Detection sensitivity using different reporter fluorochromes was tested with the LX 100, and also different assay formats were evaluated for their suitability for rapid testing. A blind laboratory trial was carried out December 22-27, 1999 to evaluate bead assays for multiplex identification of Bg and Bt PCR products. This report summarizes the assay development, fluorochrome comparisons, and the results of the blind trial conducted at LLNL for the laboratory evaluation of the LX 100 flow analyzer.
Date: May 5, 2000
Creator: Venkateswaran, K.S.; Nasarabadi, S. & Langlois, R.G.
Partner: UNT Libraries Government Documents Department

Final Report for CRADA Agreement , AL-C-2006-01 with Microsens Biotechnologies: Detection of the Abnormal Prion Protein in Blood by Improving the Extraction of this Protein

Description: Several conditions were examined to optimize the extraction protocol using Seprion beads for the abnormal prion protein. Different combinations of water, hexafluro-2-propanol and formic acid were used. The results of these extraction protocols showed that the magnetic beads coated with Seprion reagents were subject to degradation, themselves, when the extraction conditions that would solubilize the abnormal prion protein were used. These compounds caused interference in the immunoassay for the abnormal prion protein and rendered these protocols incompatible with the assay systems. In an attempt to overcome this problem, another approach was then used. The coated beads were used as an integral part of the assay platform. After washing away denaturing agents, the beads with the 'captured' abnormal prion were incubated directly in the immunoassay, followed by analysis by the capillary electrophoresis. When a capillary electrophoresis electro-kinetic separation was attempted, the beads disturbed the analysis making it impossible to interpret. A pressure separation method was then developed for capillary electrophoresis analysis. When 20 samples, 5 of which were positive were analyzed, the assay identified 4 of the 5 positives and had no false positives. When a larger number of samples were analyzed the results were not as good - there were false positives and false negatives. It was then observed that the amount of beads that were loaded was dependent upon how long the beads were allowed to settle before loading them into the capillary. This resulted in unacceptable variations in the results and explained that when large numbers of samples were evaluated the results were not consistent. Because the technical difficulties with using the Seprion beads could not be overcome at this time, another approach is underway that is outside of the scope of this CRADA. No further agreements have been developed. Because the results were not favorable, no manuscripts ...
Date: March 31, 2009
Creator: Schmerr, Mary Jo
Partner: UNT Libraries Government Documents Department

Umatilla Hatchery Satellite Facilities; Operations and Maintenance, Annual Report 2001.

Description: The Confederated Tribes of the Umatilla Indian Reservation (CTUIR) and Oregon Department of Fish and Wildlife (ODFW) are cooperating in a joint effort to enhance steelhead and re-establish salmon runs in the Umatilla River Basin. As an integral part of this program, Bonifer Pond, Minthorn Springs, Imeques C-mem-ini-kem, Thornhollow and Pendleton satellite facilities are operated for acclimation and release of juvenile summer steelhead (Oncorhynchus mykiss), fall and spring chinook salmon (O. tshawytscha) and coho salmon (O. kisutch). Minthorn is also used for holding and spawning adult summer steelhead and Three Mile Dam and South Fork Walla Walla facilities are used for holding and spawning chinook salmon. In some years, Three Mile Dam may also be used for holding and spawning coho salmon. In the spring of 2002, summer steelhead were acclimated and released at Bonifer Pond (54,917), Minthorn Springs (47,521), and Pendleton (54,366). Yearling coho (1,621,857) were also acclimated and released at Pendleton. Yearling spring chinook salmon (876,121) were acclimated and released at Imeques C-mem-ini-kem. At Thornhollow, 520,564 yearling fall chinook and 307,194 subyearling fall chinook were acclimated. In addition, 104,908 spring chinook were transported to Imeques C-mem-ini-kem in November for release in the spring of 2003. CTUIR and ODFW personnel monitored the progress of outmigration for juvenile releases at the Westland Canal juvenile facility. Nearly all juveniles released in the spring migrated downstream prior to the trap being opened in early July. A total of 100 unmarked and 10 marked summer steelhead were collected for broodstock at Three Mile Dam from September 21, 2001, through April 2, 2002. An estimated 180,955 green eggs were taken from 36 females and were transferred to Umatilla Hatchery for incubation and rearing. A total of 560 adult and 26 jack spring chinook salmon were collected for broodstock at Three Mile Dam from April ...
Date: May 1, 2003
Creator: Rowan, Gerald
Partner: UNT Libraries Government Documents Department

Heightened sense for sensing: recent advances in pathogen immunoassay sensing platforms

Description: As part of its own defense mechanism, most bacteria have developed an innate ability to enable toxic secretion to ward off potential predators or invaders. However, this naturally occurring process has been abused since over production of the bacteria's toxin molecules could render them as potential bioweapons. As these processes (also known as ''black biology'') can be clandestinely performed in a laboratory, the threat of inflicting enormous potential damage to a nation's security and economy is invariably clear and present. Thus, efficient detection of these biothreat agents in a timely and accurate manner is highly desirable. A wealth of publications describing various pathogen immuno-sensing advances has appeared over the last few years, and it is not the intent of this review article to detail each reported approach. Instead, we aim to survey a few recent highlights in hopes of providing the reader an overall sense of the breath of these sensing systems and platforms. Antigen targets are diverse and complex as they encompass proteins, whole viruses, and bacterial spores. The signaling processes for these reported immunoassays are usually based on colorimetric, optical, or electrochemical changes. Of equal interest is the type of platform in which the immunoassay can be performed. A few platforms suitable for pathogen detection are described.
Date: January 9, 2007
Creator: Fischer, N; Tarasow, T & Tok, J B
Partner: UNT Libraries Government Documents Department

Adsorption of biometals to monosodium titanate in biological environments

Description: Monosodium titanate (MST) is an inorganic sorbent/ion exchanger developed for the removal of radionuclides from nuclear wastes. We investigated the ability of MST to bind Cd(II), Hg(II), or Au(III) to establish the utility of MST for applications in environmental decontamination or medical therapy (drug delivery). Adsorption isotherms for MST were determined at pH 7-7.5 in water or phosphate-buffered saline. The extent of metal binding was determined spectroscopically by measuring the concentrations of the metals in solution before and after contact with the MST. Cytotoxic responses to MST were assessed using THP1 monocytes and succinate dehydrogenase activity. Monocytic activation by MST was assessed by TNF{alpha} secretion (ELISA) with or without lipopolysaccharide (LPS) activation. MST sorbed Cd(II), Hg(II), and Au(III) under conditions similar to that in physiological systems. MST exhibited the highest affinity for Cd(II) followed by Hg(II) and Au (III). MST (up to 100 mg/L) exhibited only minor (&lt; 25% suppression of succinate dehydrogenase) cytotoxicity and did not trigger TNF{alpha} secretion nor modulate LPS-induced TNF{alpha} secretion from monocytes. MST exhibits high affinity for biometals with no significant biological liabilities in these introductory studies. MST deserves further scrutiny as a substance with the capacity to decontaminate biological environments or deliver metals in a controlled fashion.
Date: June 6, 2005
Creator: HOBBS, D.T.; MESSER, R. L. W.; LEWIS, J. B.; CLICK, D. R. LOCKWOOD, P. E. & WATAHA, J. C.
Partner: UNT Libraries Government Documents Department

Field-Portable Immunoassay Instruments and Reagents to Measure Chelators and Mobile Forms of Uranium

Description: The goals for the 3-year project period are (1) to test and validate the present uranium sensor and develop protocols for its use at the NABIR Field Research Center; (2) to develop new reagents that will provide superior performance for the present hand-held immunosensor; and (3) to develop new antibodies that will permit this sensor to also measure other environmental contaminants (chromium, mercury, and/or DTPA). Sensor design modifications are underway via international collaborations. New reagents that will provide superior performance for the present hand-held immunosensor are being prepared and tested. New methods have been developed, to produce recombinant forms of metal-specific monoclonal antibodies for use with the sensor. Site-directed mutagenesis experiments are underway to determine the mechanisms of binding. Immunization experiments with sheep and rabbits to develop new recombinant forms of antibodies to metal-chelate complexes (chromium, mercury, and/or DTPA) have been initiated.
Date: June 1, 2003
Creator: Blake, Diane A.
Partner: UNT Libraries Government Documents Department

Field-Portable Immunoassay Instruments and Reagents to Measure Chelators and Mobile Forms of Uranium

Description: Progress Report Date: 01/23/06 (report delayed due to Hurricane Katrina) Report of results to date: The goals of this 3-year project are to: (1) update and successfully deploy our present immunosensors at DOE sites; (2) devise immunosensor-based assays for Pb(II), Hg(II), chelators, and/or Cr(III) in surface and groundwater; and (3) develop new technologies in antibody engineering that will enhance this immunosensor program. Note: Work on this project was temporarily disrupted when Hurricane Katrina shut down the University on August 29, 2005. While most of the reagents stored in our refrigerators and freezers were destroyed, all of our hybridoma cell lines were saved because they had been stored in liquid nitrogen. We set up new tissue culture reactors with the hybridomas that synthesize the anti-uranium antibodies, and are purifying new monoclonal antibodies from these culture supernatants. Both the in-line and the field-portable sensor were rescued from our labs in New Orleans in early October, and we continued experiments with these sensors in the temporary laboratory we set up in Hammond, LA at Southeastern Louisiana University.
Date: January 23, 2006
Creator: Blake, Diane A.
Partner: UNT Libraries Government Documents Department

Microfluidic System for Solution Array Based Bioassays

Description: The objective of this project is to demonstrate new enabling technology for multiplex biodetection systems that are flexible, miniaturizable, highly automated, low cost, and high performance. It builds on prior successes at LLNL with particle-based solution arrays, such as those used in the Autonomous Pathogen Detection System (APDS) successfully field deployed to multiple locations nationwide. We report the development of a multiplex solution array immunoassay based upon engineered metallic nanorod particles. Nanobarcodes{reg_sign} particles are fabricated by sequential electrodeposition of dissimilar metals within porous alumina templates, yielding optically encoded striping patterns that can be read using standard laboratory microscope optics and PC-based image processing software. The addition of self-assembled monolayer (SAM) coatings and target-specific antibodies allows each encoded class of nanorod particles to be directed against a different antigen target. A prototype assay panel directed against bacterial, viral, and soluble protein targets demonstrates simultaneous detection at sensitivities comparable to state of the art immunoassays, with minimal cross-reactivity. Studies have been performed to characterize the colloidal properties (zeta potential) of the suspended nanorod particles as a function of pH, the ionic strength of the suspending solution, and surface functionalization state. Additional studies have produced means for the non-contact manipulation of the particles, including the insertion of magnetic nickel stripes within the encoding pattern, and control via externally applied electromagnetic fields. Using the results of these studies, the novel Nanobarcodes{reg_sign} based assay was implemented in a prototype automated system with the sample processing functions and optical readout performed on a microfluidic card. The unique physical properties of the nanorod particles enable the development of integrated microfluidic systems for biodefense, protein expression studies, and other applications.
Date: February 10, 2006
Creator: Dougherty, G M; Tok, J B; Pannu, S S & Rose, K A
Partner: UNT Libraries Government Documents Department

Mass Spectrometric Immunoassay for Parathyroid Hormone Related Protein (PTHrP)

Description: Many cancers, including prostate, breast and lung express parathyroid hormone related protein (PTHrP). Despite the common tumor overexpression of PTHrP, serum levels of PTHrP are not commonly elevated in affected patients. They postulate that the reasons for the discrepancy between tissue and serum measurements of PTHrP are the inadequate sensitivity and specificity of current PTHrP serum assays. To improve the clinical value of PTHrP serum assays for the cancer patient, they are developing a new generation of novel and ultrasensitive PTHrP serum immunoassays based on immunoaffinity purification, nanospray liquid chromatography tandem mass spectrometry (LC/MS/MS) and accelerator mass spectrometry (AMS).
Date: June 16, 2000
Creator: Zheng, K.; Rivera, J.D.; Vogel, J.S.; Buchholz, B.A.; Burton, D.W.; Deftos, L.J. et al.
Partner: UNT Libraries Government Documents Department