Electrophoresis titration method for determining activity of peroxidase
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A peroxidase, active technology, applied in the field of bioanalytical technology and medical testing, can solve the problems of low throughput, high cost, large sample consumption and so on
Active Publication Date: 2017-07-21
SHANGHAI JIAO TONG UNIV
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Embodiment 1
[0079] In the present embodiment, the horseradish peroxidase activity is determined by electrophoretic titration method, and the electrophoretic titration method is as follows: figure 1 shown. First, fill the channel part of the chip with gel, inject the mixed solution a of hydrochloric acid solution and background electrolyte solution into the anode pool, and inject the mixed solution b of chemiluminescent bottom solution and background electrolyte solution into the cathode pool; The oxidase solution is injected into the cathode cell to catalyze the reaction of the chemiluminescent base solution to generate the excited state luminol anion L* that emits blue fluorescence; after the enzyme-catalyzed reaction is stable for a period of time, a voltage is applied to make the excited state generated in the cathode cell The state luminol anion L* moves to the anode and interacts with the hydrogen ion H in the anode pool moving to the cathode + MRB is formed, and MRB moves to the an...
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Abstract
The present invention discloses an electrophoresis titration method for determining the activity of peroxidase. The electrophoresis titration method comprises: filling the channel part of a chip with a gel, injecting the mixed solution a of a hydrochloric acid solution and a background electrolyte solution into an anode pool, and injecting the mixed solution b of a chemiluminescent base solution and a background electrolyte solution into a cathode pool; injecting a peroxidase solution into the cathode pool to catalyze the chemiluminescent base solution to react so as to produce excitation state luminol anions L*; after the enzyme catalytic reaction is stabilized, applying an electric field to make the excitation state luminol anions L* generated in the cathode pool and the hydrogen ions H<+> in the anode pool form MRB, wherein the generated MRB moves to the anode; and according to the functional relationship between the MRB movement rate and the activity of the peroxidase, determining the activity index of the peroxidase. With the method of the present invention, various enzyme indexes of the peroxidase can be effectively and rapidly determined without complex optical detection equipment.
Description
technical field [0001] The invention relates to the field of biological analysis technology and medical inspection, in particular to an electrophoretic titration method for measuring peroxidase activity. Background technique [0002] Enzyme analysis in clinical medicine (Abraham, E.C.; Huff, T.A.; Cope, N.D.; Wilson, J.B.; Bramsome ELISA,, E.D.Jr.; Huisman, T.H.J. Diabetes 1978, 27, 931–937.) Biology (Tannu, N.S.; Hemby , S.E.Nature Protocols 2006,1,1732-1942.) and drug screening (Li, Juan; Wu, Ling-Jie; Guo, Shan-Shan; Fu, Hua-E; Chen, Guo-Nan; Yang, Huang-Hao ,NANOSCALE,2013,5,619-623.) plays an important role in the field. Traditional detection methods are generally based on the detection of substrate reduction and product formation in the enzyme reaction system to determine the catalytic rate, activity, and Michaelis constant. At present, many methods can be used for enzyme detection, such as mass spectrometry (Northen, TR; Lee, JC; Hoang, L; Raymond, J; Hwang, DR; Yan...
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