Dual Stage Microvalue and Method of Use

A micro-machined drug delivery device and method of use for the delivery of labile drugs is disclosed. A micro-machined sensing device and method of use is also disclosed. A micro-machined drug delivery and sensing device and method of use is additionally disclosed. All three devices are intended to be inserted into a patient\u27s body. The drug delivery devices allow for the mixing of drug components prior to the release of the mixture into the patient\u27s body where the mixture is labile. The micro-machined sensing device is suitable for monitoring the concentration of a specific chemical in a patient\u27s body fluids when the monitoring requires a labile reagent that must be mixed prior to introduction of the body fluid into the sensing device. The micro-machined drug delivery and sensing device is especially applicable in situations where the prompt delivery of labile drugs is necessary

Metal-Resistance Genetically Engineered Bacteria

Bacterial-based electrochemical and optical sensing systems that respond in a highly selective and sensitive manner to antimonite and arsenite have been developed. This was accomplished by using genetically engineered bacteria bearing one of two plasmids constructed for our studies. The first plasmid, pBGD23, contains the operator/promoter region (O/P) and the gene of the ArsR protein from the ars operon upstream from the β-galactosidase gene. In the absence of antimonite/arsenite, ArsR binds to the 0/P site and prevents the transcription of the genes for ArsR and β-galactosidase, thus blocking expression of these proteins. When antimonite or arsenite is present in the sample, it binds to the ArsR protein, causing a conformational change in ArsR that leads to its release from the O/P site of the plasmid, thus allowing for the expression of β-galactosidase. Then, the amount of β-galactosidase expressed is quantified by using a substrate that produces a product that can be monitored electrochemically. In the second plasmid, pRLUX, the gene for ArsR is upstream from the reporter gene, luxAB, that encodes for the enzyme luciferase, whose activity can be monitored by bioluminescence. These bacterial sensing systems have excellent detection limits, respond selectively to arsenite and antimonite, and show no significant response to phosphate, sulfate, nitrate, and carbonate

Polypeptides, Systems, and Methods Useful for Detecting Glucose

The presently-disclosed subject matter provides biosensors for detecting molecules of interest. The biosensors include a polypeptide capable of selectively-binding glucose, wherein the polypeptide molecule is selected from: an unnatural analogue of wild type glucose binding protein; a fragment of wild type glucose binding protein; and an unnatural analogue fragment of wild type glucose binding protein

Aequorin and Obelin Mutants with Differing Wavelengths and Bioluminescence

The invention relates to aequorin and obelin mutants whose emission is shifted with respect to wild type. The shift in emission is accomplished using a combination of mutations of amino acids within aequorin or obelin that affect bioluminescence; use of different types of chromophores, i.e., coelenterazines with variable emission characteristics; and modifications of the photoprotein with fluorophores that will allow for emission of light at longer wavelengths as a result of energy transfer. Additionally, an assay employing aequorin mutants to test for HIV-1 protease inhibitors is disclosed

Methods and Kit for Determination of Prostacyclin in Plasma

A solid-phase immunoassay for 6-keto-Prostaglandin F1α, the stable hydrolysis product of prostacyclin (Prostaglandin I2) is disclosed. Prostacyclin, a potent vasodilator with anti-platelet and anti-proliferative properties is an effective treatment for primary pulmonary hypertension and pulmonary arterial hypertension associated with scleroderma and scleroderma-like syndrome. Levels of 6-keto-Prostaglandin F1α can be directly correlated with levels of prostacyclin. Therefore, 6-keto-Prostaglandin F1α has become the indicator of choice to measure prostacyclin levels. The single step immunoassay for 6-keto-Prostaglandin F1α uses the bioluminescent protein, aequorin as a label. Analyte-label conjugates were constructed by linking the carboxyl group of 6-keto-Prostaglandin F1α and lysine residues of aequorin by chemical conjugation methods. The binding properties of 6-keto-Prostaglandin F1α towards its antibody and the bioluminescent properties of aequorin are retained in the conjugate. The concentration of 6-keto-Prostaglandin F1α after extraction from plasma shows good correlation with the concentration of 6-keto-Prostaglandin F1α obtained without prior extraction of the same plasma sample. The assay allows the measurement of 6-keto-Prostaglandin F1α directly in plasma without any pre-treatment of the samples, which results in a much simpler method with a faster assay time

TLR4 Signaling is Involved in Brain Vascular Toxicity of PCB153 Bound to Nanoparticles

PCBs bind to environmental particles; however, potential toxicity exhibited by such complexes is not well understood. The aim of the present study is to study the hypothesis that assembling onto nanoparticles can influence the PCB153-induced brain endothelial toxicity via interaction with the toll-like receptor 4 (TLR4). To address this hypothesis, TLR4-deficient and wild type control mice (males, 10 week old) were exposed to PCB153 (5 ng/g body weight) bound to chemically inert silica nanoparticles (PCB153-NPs), PCB153 alone, silica nanoparticles (NPs; diameter, 20 nm), or vehicle. Selected animals were also subjected to 40 min ischemia, followed by a 24 h reperfusion. As compared to exposure to PCB153 alone, treatment with PCB153-NP potentiated the brain infarct volume in control mice. Importantly, this effect was attenuated in TLR4-deficient mice. Similarly, PCB153-NP-induced proinflammatory responses and disruption of tight junction integrity were less pronounced in TLR4-deficient mice as compared to control animals. Additional in vitro experiments revealed that TLR4 mediates toxicity of PCB153-NP via recruitment of tumor necrosis factor-associated factor 6 (TRAF6). The results of current study indicate that binding to seemingly inert nanoparticles increase cerebrovascular toxicity of PCBs and suggest that targeting the TLR4/TRAF6 signaling may protect against these effects

Stimuli-Responsive Hydrogel Microdomes Integrated with Genetically Engineered Proteins for High-Throughput Screening of Pharmaceuticals

A hydrogel microdome that can swell in response to a stimuli or target molecule is formed by polymerizing a mixture comprising a monomer capable of forming a hydrogel with a biopolymer. An array of hydrogel microdomes can be formed on a substrate by microspotting the mixture and polymerizing. The array can be used for high-throughput screening of analytes as well as for use as an actuator and biosensor using the swelling property of the hydrogel

Homogeneous enzyme-linked competitive binding assay for biotin based on the avidin-biotin interaction

A homogeneous enzyme-linked competitive-binding assay for biotin with glucose-6-phosphate dehydrogenase (G6PDH), is described. This assay is based on the interaction between a G6PDH/biotin conjugate with avidin, a natural binder for biotin. In the absence of biotin in the assay mixture, this interaction results in 100% inhibition of the enzyme conjugate. In the presence of biotin, the enzymatic activity of the conjugate is regained in an amount related to the concentration of the vitamin in the sample. Extremely steep, gate-like dose/response curves, attributable to the relative binding affinities of avidin for biotin and the conjugate, are observed. The detection limits of the system vary with the amounts of avidin and enzyme/biotin conjugate used. The method is rapid and sensitive and is evaluated for the direct determination of biotin in vitamin tablets.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/27454/1/0000494.pd

Systems and Methods for Diagnosis and Monitoring of Bacteria-Related Conditions

The presently-disclosed subject matter provides systems, methods, and kits for diagnosing and/or monitoring a bacteria-related condition of interest in a subject by providing a cell sensing system, each system containing a reporter molecule capable of detecting binding of a quorum sensing molecule and capable of generating a detectable signal