Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.3.1 (alkaline phosphatase)
47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Although surface charge has been shown to affect the adhesion and morphology of a variety of cell types, the interactions of bone marrow stromal cells with charged surfaces still remain unclear. A novel electrical stimulation system was used to investigate the interactions between rat bone marrow stromal cells and charged substrates in this study. A conductive and transparent indium tin oxide (ITO) coating was used as an electret substrate. Rat marrow stromal cells were cultured on positive, negative, and uncharged ITO surfaces. Cell attachment, morphology, alkaline phosphatase activity, and expression of osteopontin and collagen type III were assessed using histochemical staining, immunolabeling, and fluorescence microscopy. Voltages of 0.7, 0.8, 0.9, and 1.0 V applied to the substrates created surface potentials but were insufficient to decompose the media. On positively charged ITO, cell attachment was enhanced in serum-supplemented and serum-free media. Furthermore, decreases in cell spreading, alkaline phosphatase activity, and osteopontin were observed in cells grown on the positively charged ITO. These data indicate that positively charged surfaces enhance cell attachment but suppress cell spreading and differentiation of rat marrow stromal cells.
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PMID:Attachment, morphology, and protein expression of rat marrow stromal cells cultured on charged substrate surfaces. 974 14

Changes in hematological and serum biochemistry parameters in female zinc (Zn)-dosed farm-raised mallards (Anas platyrhynchos) fed four different diets were examined. Sixty ducks received an average dose of 0.97 g of Zn in the form of eight, 3.30-mm diameter shot pellets containing 98% Zn and 2% tin, and another 60 ducks were sham-dosed as controls. Fifteen ducks from each of the two dosing groups were assigned to one of four dietary treatments: corn only, corn with soil, commercial duck ration only, or commercial duck ration with soil. Shot-pellet dissolution rates ranged from 7 mg/Zn/day to 27 mg/Zn/day. Regardless of diet, the Zn dose resulted in mortality; incoordination; paralysis and anorexia; decreased body, liver, pancreas, gonad, and gizzard weight; increased kidney weight; and macroscopic lesions. Zn-dosed ducks had a lower mean erythrocyte packed cell volume (PCV), higher mean reticulocyte count, and a greater number of individuals with immature and/or abnormal erythrocytes, than did control mallards. Mean total leucocyte counts were higher in Zn-dosed ducks than in controls. Zn-dosed ducks that had soil available had higher leucocyte counts than those without soil. Zn-dosed ducks were characterized by a marked heterophilia and relative lymphopenia. In Zn-dosed ducks, the mean lymphocyte count was highest in those provided a commercial duck ration, and lowest in those fed corn. In control ducks, the mean lymphocyte count was highest in ducks fed corn, and lowest in those provided soil along with a commercial duck ration. Zn-dosed mallards had higher serum aspartate aminotransferase and amylase levels, and lower alkaline phosphatase activities than control ducks. Serum phosphorus and uric acid concentrations were higher, and calcium, glucose, and total protein levels lower, in Zn-dosed ducks than in control ducks. Diet did affect serum calcium, phosphorus, total protein, and uric acid concentrations. Differences in erythrocyte and leucocyte parameters, serum enzyme activities, and metabolite concentrations were associated with dose and diet effects. Diets high in protein and other organic matter and calcium and phosphorus did not prevent or substantially alleviate Zn toxicosis in farm-raised mallard ducks.
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PMID:Influence of diet on the hematology and serum biochemistry of zinc-intoxicated mallards. 1068 52

Impedance biosensor chips were developed for detection of Escherichia coli O157:H7 based on the surface immobilization of affinity-purified antibodies onto indium tin oxide (ITO) electrode chips. The immobilization of antibodies onto ITO chips was carried out using an epoxysilane monolayer to serve as a template for chemical anchoring of antibodies. The surface characteristics of chips before and after the binding reaction between the antibodies and antigens were characterized by atomic force microscopy (AFM). The patterns of the epoxysilanes monolayer, antibodies, and E. coli cells were clearly observed from the AFM images. Alkaline phosphatase as the labeled enzyme to anti-E. coli O157:H7 antibody was used to amplify the binding reaction of antibody-antigen on the chips. The biocatalyzed precipitation of 5-bromo-4-chloro-3-indolyl phosphate by alkaline phosphatase on the chips in pH 10 PBS buffer containing 0.1 M MgCl2 increased the electron-transfer resistance for a redox probe of Fe(CN)6(3-/4-) at the electrode-solution interface or the electrode resistance itself. Electrochemical impedance spectroscopy and cyclic voltammetric method were employed to follow the stepwise assembly of the systems and the electronic transduction for the detection of E. coli. The biosensor could detect the target bacteria with a detection limit of 6 x 10(3) cells/mL. A linear response in the electron-transfer resistance for the concentration of E. coli cells was found between 6 x 10(4) and 6 x 10(7) cells/mL.
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PMID:Immunobiosensor chips for detection of Escherichia coil O157:H7 using electrochemical impedance spectroscopy. 1234 88

This paper describes immobilization of DNA onto the interior walls of poly(dimethylsiloxane) (PDMS) microsystems and its application to an enzyme-amplified electrochemical DNA assay. DNA immobilization was carried out by silanization of the PDMS surface with 3-mercaptopropyltrimethoxysilane to yield a thiol-terminated surface. 5'-acrylamide-modified DNA reacts with the pendant thiol groups to yield DNA-modified PDMS. Surface-immobilized DNA oligos serve as capture probes for target DNA. Biotin-labeled target DNA hybridizes to the PDMS-immobilized capture DNA, and subsequent introduction of alkaline phosphatase (AP) conjugated to streptavidin results in attachment of the enzyme to hybridized DNA. Electrochemical detection of DNA hybridization benefits from enzyme amplification. Specifically, AP converts electroinactive p-aminophenyl phosphate to electroactive p-aminophenol, which is detected using an indium tin oxide interdigitated array (IDA) electrode. The IDA electrode eliminates the need for a reference electrode and provides a steady-state current that is related to the concentration of hybridized DNA. At present, the limit of detection of the DNA target is 1 nM in a volume of 20 nL, which corresponds to 20 attomoles of DNA.
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PMID:Immobilization of DNA onto poly(dimethylsiloxane) surfaces and application to a microelectrochemical enzyme-amplified DNA hybridization assay. 1645 8

Signal amplification and noise reduction are crucial for obtaining low detection limits in biosensors. Here, we present an electrochemical immunosensor in which the signal amplification is achieved using p-aminophenol (AP) redox cycling by hydrazine, and the noise level is reduced by implementing a low background current. The redox cycling is obtained in a simple one-electrode, one-enzyme format. In a sandwich-type heterogeneous immunosensor for mouse IgG, an alkaline phosphatase label converts p-aminophenyl phosphate into AP for 10 min. This generated AP is electrooxidized at an indium tin oxide (ITO) electrode modified with a partially ferrocenyl-tethered dendrimer (Fc-D). The oxidized product, p-quinone imine (QI), is reduced back to AP by hydrazine, and then AP is electrooxidized again to QI, resulting in redox cycling. Moreover, hydrazine protects AP from oxidation by air, enabling long incubation times. The small amount of ferrocene in a 0.5% Fc-D-modified ITO electrode, where 0.5% represents the ratio of ferrocene groups to dendrimer amines, results in a low background current, and this electrode exhibits high electron-mediating activity for AP oxidation. Moreover, there is insignificant hydrazine electrooxidation on this electrode, which also results in a low background current. The detection limit of the immunosensor using a 0.5% Fc-D-modified electrode is 2 orders of magnitude lower than that of a 20% Fc-D-modified electrode (10 pg/mL vs 1 ng/mL). Furthermore, the presence of hydrazine reduces the detection limit by an additional 2 orders of magnitude (100 fg/mL vs 10 pg/mL). These results indicate that the occurrence of redox cycling combined with a low background current yields an electrochemical immunosensor with a very low detection limit (100 fg/mL). Mouse IgG could be detected at concentrations ranging from 100 fg/mL to 100 microg/mL (i.e., 9 orders of magnitude) in a single assay.
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PMID:Electrochemical immunosensor using p-aminophenol redox cycling by hydrazine combined with a low background current. 1731 7

A chip with integrated electrophoretic and electrochemical systems was developed to manipulate either an individual microbead or a cell inside a microwell electrode (MWE) for electrochemical measurement. The optimal MWE geometry (30 microm diameter and 25 microm depth) was designed to accommodate the micro particles according to the simulated results. A chip device was sequentially built from a slide patterned with Pt electrodes, an adhesive tape defined with a flow channel (200 microm in width and 25 microm in height), and an indium tin oxide (ITO) cover. The MWE not only generated an active electrophoretic force to trap the particle but also provided a low flow velocity area (LFVA) to stabilize the trapped bead or cell in a continuous flow. Scanning electrochemical microscopy (SECM) theory was employed to explain the electrochemical behaviors of the MWE. An enhanced current was confirmed as the redox recycling effect on the conductive ITO cover. The catalytic reaction of an individual alkaline phosphatase coated microbead (ALP-bead) was electrochemically detected with the MWE after being trapped. The ALP on the trapped ALP-bead catalyzed the hydrolysis of p-aminophenylphosphate (PAPP) to p-aminophenol (PAP), and then a decaying amperogram (+0.3 V vs. Ag/AgCl) due to a tiny PAP quantity around the MWE was observed.
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PMID:Entrapment and measurement of a biologically functionalized microbead with a microwell electrode. 1937 Feb 35

Scanning electrochemical microscopy (SECM) was used for the analysis of single-cell gene-expression signals on the basis of a reporter system. We microfabricated a single-cell array on an Indium tin oxide (ITO) electrode comprising 4 x 4 SU-8 microwells with a diameter of 30microm and a depth of 25microm. HeLa cells transfected with plasmid vectors encoding the secreted alkaline phosphatase (SEAP) were seeded in the microwell at a concentration of 1 cell per well by positive-dielectrophoresis (pDEP). A pDEP pulse of 3.0Vpp and 1MHz was applied between the microwell array/ITO electrode and an ITO counter electrode located on the top of the flow-cell assembly of the microdevice. The electrochemical responses of the individual HeLa cells transfected with SEAP were significantly larger than those of the wild-type HeLa cells. The electrochemical response of the transfected single cells was statistically distinguishable from that of wild-type HeLa cells. The size of the wells and the material of the single-cell array were optimized in order to evaluate the tumor necrosis factor alpha (TNF-alpha)-induced activation process of nuclear factor kappa B (NFkappaB) that was used as the model for on-chip monitoring of cellular signal transduction.
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PMID:Electrochemical single-cell gene-expression assay combining dielectrophoretic manipulation with secreted alkaline phosphatase reporter system. 1977 81

Cryptosporidium parvum is one of the most important biological contaminants in drinking water and generates significant risks to public health. Due to low infectious dose of C. parvum, remarkably sensitive detection methods are required for water and food industry analysis. This present study describes a simple, sensitive, enzyme amplified sandwich form of an electrochemical immunosensor using dual labeled gold nanoparticles (alkaline phosphatase and anti-oocysts monoclonal antibody) in indium tin oxide (ITO) as an electrode to detect C. parvum. The biosensor was fabricated by immobilizing the anti-oocysts McAb on a gold nanoparticle functionalized ITO electrode, followed by the corresponding capture of analytes and dual labeled gold nanoparticle probe to detect the C. parvum target. The outcome shows the sensitivity of electrochemical immune sensor enhanced by gold nanoparticles with a limit of detection of 3 oocysts/mL in a minimal processing period. Our results demonstrated the sensitivity of the new approach compared to the customary method and the immunosensors showed acceptable precision, reproducibility, stability, and could be readily applied to multi analyte determination for environmental monitoring.
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PMID:Development of electrochemical based sandwich enzyme linked immunosensor for Cryptosporidium parvum detection in drinking water. 2189 77

This paper reports chemical-chemical (CC) and electrochemical-chemical-chemical (ECC) redox cycling, for use in ultrasensitive biosensor applications. A triple chemical amplification approach using an enzymatic reaction, CC redox cycling, and ECC redox cycling is applied toward electrochemical immunosensors of cardiac troponin I. An enzymatic reaction, in which alkaline phosphatase converts 4-aminophenyl phosphate to 4-aminophenol (AP), triggers CC redox cycling in the presence of an oxidant and a reductant, and electrochemical signals are measured with ECC redox cycling after an incubation period of time in an air-saturated solution. To obtain high, selective, and reproducible redox cycling without using redox enzymes, two redox reactions [the reaction between AP and the oxidant and the reaction between the oxidized form of AP (4-quinone imine, QI) and the reductant] should be fast, but an unwanted reaction between the oxidant and reductant should be very slow. Because species that undergo outer-sphere reactions (OSR-philic species) react slowly with species that undergo inner-sphere reactions (ISR-philic species), highly OSR-philic Ru(NH(3))(6)(3+) and highly ISR-philic tris(2-carboxyethyl)phosphine (TCEP) are chosen as the oxidant and reductant, respectively. The OSR- and ISR-philic QI/AP couple allows fast redox reactions with both the OSR-philic Ru(NH(3))(6)(3+) and the ISR-philic TCEP. Highly OSR-philic indium-tin oxide (ITO) electrodes minimize unwanted electrochemical reactions with highly ISR-philic species. Although the formal potential of the Ru(NH(3))(6)(3+)/Ru(NH(3))(6)(2+) couple is lower than that of the QI/AP couple, the endergonic reaction between Ru(NH(3))(6)(3+) and AP is driven by the highly exergonic reaction between TCEP and QI (via a coupled reaction mechanism). Overall, the "outer-sphere to inner-sphere" redox cycling in the order of highly OSR-philic ITO, highly OSR-philic Ru(NH(3))(6)(3+)/Ru(NH(3))(6)(2+) couple, OSR- and ISR-philic QI/AP couple, and highly ISR-philic TCEP allows high, selective, and reproducible signal amplification. The electrochemical data obtained by chronocoulometry permit a lower detection limits than those obtained by cyclic voltammetry. The detection limit of an immunosensor for troponin I in serum, calculated from the anodic charges in chronocoulometry, is ca. 10 fg/mL.
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PMID:"Outer-sphere to inner-sphere" redox cycling for ultrasensitive immunosensors. 2220 64

Polypyrrole (PPy), as an electrical conductive polymer, has been widely investigated in biomedical fields. In this study, PPy membrane at nanoscale was electrically deposited on indium-tin oxide glass slide with sodium p-toluenesulfonate as supporting electrolyte. Electropolymerization of PPy was performed under a constant 800 mV voltage for 10 seconds. Chemical compositions and morphology were characterized by Fourier transform infrared spectroscopy (FTIR) and scanning electron microscopy (SEM). The results showed that the nanoscaled PPy particles distributed uniformly and the average diameter of PPy particles was 62 nm. Since bone cells can respond to both electrical and mechanical stimulation in vivo, pre-osteoblasts MC3T3-E1 cells were cultured ort nanostructured PPy membrane under the combined electrical and mechanical stimulation. The nano-PPy membrane was conducive to transferring uniform electrical stimulation and applying steady mechanical stimulation. It is suggested that the combined stimulation did not affect cells morphologies significantly. However, cell proliferation tested by MTT, alkaline phosphatase activities, and gene expression of Collagen-I indicated that combined stimulation can enhance the proliferation and differentiation of MC3T3-E1 cells more efficiently than single electrical stimulation or single mechanical stimulation. The combined stimulation through a nano-PPy membrane may provide a highly potential stimulated method in bone tissue engineering.
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PMID:Increased proliferation and differentiation of pre-osteoblasts MC3T3-E1 cells on nanostructured polypyrrole membrane under combined electrical and mechanical stimulation. 2398 May 1


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