51 results about "Diaphorase" patented technology

There is provided a fuel cell whose current density and maintenance ratio can be improved when at least glucose dehydrogenase and diaphorase are immobilized on an anode using an immobilizing material composed of poly-L-lysine and glutaraldehyde. The fuel cell has a structure in which a cathode 2 and an anode 1 face each other with an electrolyte layer 3 therebetween, the anode 1 being obtained by immobilizing at least glucose dehydrogenase and diaphorase on an electrode using an immobilizing material composed of poly-L-lysine and glutaraldehyde, wherein the mass ratio of the poly-L-lysine to the glutaraldehyde in an immobilizing material is 5:1 to 80:1, the mass ratio of the glucose dehydrogenase to the diaphorase is 1:3 to 200:1, and the average molecular weight of the poly-L-lysine is 21500 or more.

A fuel cell capable of directly abstracting electric power from polysaccharides, such as starch. Fuel electrode (1) is produced by fixing on electrode of carbon, etc. (11) by means of a fixative an enzyme participating in decomposition of polysaccharides to monosaccharides, an enzyme participating in decomposition of formed monosaccharides, a coenzyme (e.g., NAD<+>, NADP<+>, etc.) whose reduced form is produced in accordance with an oxidation reaction during the process of monosaccharide decomposition, a coenzyme oxidase (e.g., diaphorase) capable of oxidizing the reduced form of coenzyme (e.g., NADH, NADPH, etc.), and an electron mediator (e.g., ACNQ, vitamin K3, etc.) capable of receiving electrons generated by the coenzyme oxidation from the coenzyme oxidase and delivering the same to the electrode (11). The fuel electrode (1) is disposed opposite to air electrode (5) through electrolyte layer (3) to thereby construct a fuel cell.

The present invention relates to a dairy cow sub-clinical ketosis diagnosis test paper which belongs to the art of the veterinary clinical laboratory science. The present invention comprises a support cushion and a test cushion that is fixed on the support cushion; and the present invention is characterized in that the test cushion of the test paper comprises the following reagents: D-(-)-3-Hydroxybutyrate Dehydrogenase with 2270U/mg and 50mg-100mg; nicotinamide adenine dinucleotide with 300mg-500mg; diaphorase with 35U/mg-45U/mg and 500-1,000mg; NBT with 100mg-200mg; Tris Base with molecular weight 121.14 and 1.21g; oxalic acid with molecular 126 and 126mg; fucose with molecular 383 and 4-6g; and distilled water with 100ml. The dairy cow sub-clinical ketosis diagnosis test paper of the present invention is used conveniently and has the advantages of economic practicality, high sensitivity and specificity, and good stability. The test paper is used to test D-(-)-3-Hydroxybutyrate and D-(-)-3-Hydroxybutyrate sodium in the milk of the milch cow.

The invention discloses a method for detecting TMAO (trimethylamine oxide) by an enzyme method and application thereof. The detection method utilizes a multi-enzyme combination system containing TMAOTDM (demethylase), FADH (formaldehyde dehydrogenase), FDH (formate dehydrogenase) and diaphorase, so as to perform fluorescence determining on the TMAO; the method can be used for developing a TMAO detection kit. The method for detecting the TMAO by the enzyme method has the advantages that the defect of failure to directly determine based on dehydrogenase and diaphorase coupling because of no participation of NAD dependence dehydrogenase in the metabolism path of the TMAO is overcome; by adopting the multi-enzyme coupling of TDM-FADH-FDH-diaphorase, the system for fluorescence determining ofthe TMAO is constructed, and one TMAO molecule is decomposed into two fluorescence signals; the sensitivity is high, the detection is rapid and stable, the repeatability is good, the operation is simple and convenient, and the cost is lower.

The invention relates to diagnosis test paper for diabetic ketosis and other symptoms of relatively high ketone body. The diagnosis test paper comprises the following reagents: (1) first-phase immersion liquid which comprises a stabilizer and a protective agent, bovine serum albumin, oxalic acid, NAD<+>, NADP<+>, diaphorase, 3-hydroxybutyrate dehydrogenase and buffer solution; and (2) second-phase immersion liquid which comprises NBT, PMS and an organic solvent. The diagnosis test paper is prepared by the following steps of: soaking filter paper in the first-phase immersion liquid, sucking redundant immersion liquid and quickly and warmly drying the soaked filter paper at the temperature of between 20 and 50 DEG C; soaking the dried filter paper in the second-phase immersion liquid and drying at the temperature of between 20 and 50 DEG C again to obtain base paper of the diagnosis test paper; and sticking the base paper on a plastic base and cutting into pieces to obtain the diagnosisreagent strips. The diagnosis test paper can be assembled into various detection devices, has the advantages of convenient operation, fast detection, good sensitivity, high accuracy and the like, cansemiquantitatively detect the 3-hydroxybutyric acid content of milk, urine and blood, and has good application prospect.

Disclosed is an enzyme electrode comprising an electrode system comprising an insulating substrate having formed thereon a working electrode, a counter electrode and optionally a reference electrode, and a reaction layer formed on the electrode system, wherein the reaction layer comprises diaphorase (DI), 12α-hydroxysteroid dehydrogenase (12α-HSD) and nicotinamide adenine dinucleotide synthetase (NADS), at least a portion of the reaction layer being superposed on the working electrode, wherein the DI, 12α-HSD and NADS contained in the reaction layer are immobilized on the surface of the working electrode, so that a compound generated in the reaction layer can reach the surface of the working electrode. Also disclosed are a method for and a system for determining the concentration of adenosine triphosphate (ATP) in a sample by using the above-mentioned enzyme electrode.

The invention relates to a fluorescent probe molecule for detecting diaphorase based on benzopyranidonitrile, a preparation method and an application thereof. The molecular structure of the fluorescent probe is shown below. Further disclosed are a preparation method of the fluorescent probe molecule for detecting the diaphorase based on benzopyranidonitrile and the application of detecting the diaphorase in various substances, such as water samples and cells.

The present invention provides a gene derived from a thermophilic Bacillus, comprising the nucleotide sequence of SEQ ID No.2 and encoding a thermostable diaphorase comprising the amino acid sequence of SEQ ID No.1, a recombinant vector possessing the gene, a transformant with the recombinant vector and a process for producing the thermostable diaphorase by using the transformant.

The invention provides a diaphorase mutant with high thermal stability, a diaphorase mutant gene with high thermal stability and a preparation method of the diaphorase mutant. The diaphorase mutant isobtained through mutation of the 122nd position amino acid in the amino acid sequence of wild-type diaphorase from glycine to aspartic acid, and the amino acid sequence of the diaphorase mutant is shown in SEQ ID NO: 1. The enzymatic properties of the diaphorase mutant are the same as the enzymatic properties of the wild-type diaphorase, the thermal stability of the diaphorase mutant is significantly higher than that of the wild-type diaphorase, and the stable range of the pH value is wide; an engineering means is adopted by the preparation method of the diaphorase mutant, genes of the diaphorase mutant are cloned into an expression vector, prokaryotic expression is achieved through transformation of host cells, and a large number of highly stable diaphorase mutants can be obtained; and alot of raw materials are provided for development of clinical diagnostic reagents, and the production cost is reduced.

An enzyme-immobilized electrode is provided and includes an electrode composed of porous carbon or the like, a phospholipid layer on the electrode (11), and enzymes immobilized onto the phospholipid layer. The enzymes are, for example, diaphorase and glucose dehydrogenase. An intermediate layer composed of a protein or the like may be provided between the electrode and the phospholipid layer. By using the enzyme-immobilized electrode as a negative electrode or a positive electrode in a fuel cell using an enzyme, one or a plurality of types of enzymes can be immobilized at optimal positions onthe electrode, and thus, there are provided a highly efficient enzyme-immobilized electrode and a highly efficient fuel cell using the enzyme-immobilized electrode.

The invention discloses a detection reagent and test paper for beta-hydroxybutyrate. The detection reagent contains 3-5 KU/L of diaphorase, 9-13 KU/L of beta-hydroxybutyrate dehydrogenase, 1.2-2.5 KU/L of NAD and 0.6-1.0 KU/L of NBT. The detection reagent contains various specific enzymes for specific proportioning, thereby being capable of realizing rapid detection and more accurate detection. The test paper comprises a reaction base layer, a reaction layer, a blood filter layer and a hydrophilic layer which are stacked to form a siphon system, and rapid detection and more accurate detection are further guaranteed.

A sensor for detecting lactate dehydrogenase, used for detecting a lactate dehydrogenase content of a biological sample, comprising a substrate, an electrode group, a hydrophobic insulating layer, a cover and a reaction drug membrane. The electrode group includes a working electrode and a reference electrode and is located on the substrate, the hydrophobic insulating layer is arranged on the substrate and includes a gap, the cover covers the gap and is connected to the substrate A sample channel is formed between them to accommodate the reaction drug film, the reaction drug film is selected from the group consisting of lactic acid, nicotinamide adenine dinucleotide, diaphorase and potassium ferricyanide. The reaction drug film is in contact with the lactate dehydrogenase in the biological sample to generate a bioelectrochemical reaction, so that the electrode group undergoes an electrical signal change reflecting the content of the lactate dehydrogenase.

The invention relates to a triglyceride detection reagent, detection test paper and a test paper preparation method. The detection reagent is prepared from 0.5 to 1.5 KU/ml of lipoprotein lipase, 0.3to 0.5 KU/ml of diaphorase, 0.32 to 0.94 KU/ml of glycerol dehydrogenase, 5 to 15mmol/L of coenzyme I and 2 to 6mmol/L of a chromogenic substance, and further comprises a TritonX-100 solution with themass fraction being 0.2%-1.0% and a buffer solution with the concentration being 0.01 mol/L to 0.1 mol/L; wherein the chromogenic substance is one of nitro tetrazole blue chloride, dimethyl thiazolediphenyl tetrazole bromine blue, sodium dichlorophenol indophenolate, TNBT and iodonitrotrtrazolium chloride, the detection test paper comprises a reaction bottom layer, a reaction layer, a blood filtering layer, a capillary sample introduction layer and a hydrophilic layer, and the detection reagent is located on the upper surface of the reaction layer. The interference influence of other substances on the test result is small, and the reagent detection is more stable.

The invention discloses a method for detecting D-2-hydroxyglutarate by using FAD dependent D-2-hydroxyglutarate dehydrogenase and resazurin, and relates to the technical field of enzymatic detection.The method uses FAD dependent D-2-hydroxyglutarate dehydrogenase, resazurin and a PBS buffer solution to realize detection of D-2-hydroxyglutarate; and the FAD dependent D-2-hydroxyglutarate dehydrogenase is an AX-F-D2HGDH protein or a P-D2HGDH protein. The method is simpler in composition, lower in cost and simpler and more convenient to operate. Since no cofactor, coenzyme or diaphorase needs tobe added, introduced interference is less, sensitivity is higher, only one enzyme needs to be prepared, no exogenous cofactor NAD and the like need to be added, and meanwhile, the buffer solution canbe a PBS buffer solution which is lower in cost and easy to obtain, so that cost is lower.

The present invention relates to a method for rapidly determining the L-glutamic acid in food. The method comprises: grinding or mincing a solid or semi-solid sample by using a suitable tool, weighing a proper amount of the grinded or minced sample, adding an appropriate solvent, homogenizing, carrying out centrifuged separation, and filtering to obtain a free L-glutamic acid extraction liquid, or carrying out protein hydrolysis to obtain the L-glutamic acid extraction liquid constituting the protein structure (the liquid sample is directly filtered without the pretreatment); properly diluting the sample extraction liquid, and adjusting the pH value to 8.6; reducing nicotinamide adenine dinucleotide (NAD<+>) into nicotinamide adenine dinucleotide (NADH) in the presence of glutamate dehydrogenase(GIDH) through the L-glutamic acid of the sample detection liquid, and carrying out a reaction on the NADH and iodonitrotrtrazolium chloride (INT) under the effect of diaphorase to generate formazan; and determining the absorption peak of the formazan at 490 nm by using a light-absorption microplate reader, and calculating the L-glutamic acid content in the sample according to the absorption value of the formazan by using an external standard method. According to the present invention, the detection method has characteristics of simple operation, good accuracy, fastness, high throughput, easy popularization, and the like.