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Enzyme-linked immuno kit and method for detecting bovine lactoferrin by using same

An enzyme-linked immunosorbent reagent and bovine lactoferrin technology, which is applied in measuring devices, color/spectral characteristic measurement, instruments, etc., can solve the problems of complex operation and high detection cost, and achieve the effect of simple operation, high sensitivity and low detection cost

Inactive Publication Date: 2010-03-10
黑龙江省乳品工业技术开发中心
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The purpose of the present invention is to provide a kind of enzyme-linked immunosorbent immunoassay kit and the method for detecting bovine lactoferrin with the enzyme-linked immunosorbent immunoassay kit in order to solve the detection cost that existing bovine lactoferrin detection method exists is higher, the problem of complicated operation method

Method used

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specific Embodiment approach 1

[0010] Specific embodiment 1: The enzyme-linked immunosorbent assay kit in this embodiment includes an enzyme-linked plate, bovine lactoferrin standard, sample diluent, detection solution, substrate solution, washing solution and stop solution; wherein, the enzyme-linked plate is a 96-well enzyme Linked plate; the concentration of NaCl in the sample diluent is 150mmol / L, KH 2 PO 4 The concentration is 1.6mmol / L, Na 2 HPO 4 The concentration of KCl is 9.0mmol / L, the concentration of KCl is 2.6mmol / L, and the pH value of the sample diluent is 7.4; the detection solution is HRP-labeled mouse anti-bovine lactoferrin IgG; Cycloazine solution; the concentration of NaCl in the washing liquid is 150mmol / L, KH 2 PO 4 The concentration is 1.6mmol / L, Na 2 HPO 4 The concentration of KCl is 9.0mmol / L, the concentration of KCl is 2.6mmol / L, the mass concentration of Tween 20 is 1.25%, the pH value of washing solution is 7.4; The concentration of EDTA in the stop solution is 0.1mmol / L ...

specific Embodiment approach 2

[0012] Embodiment 2: The difference between this embodiment and Embodiment 1 is that the ELISA kit of this embodiment also includes rabbit anti-bovine lactoferrin serum. Others are the same as in the first embodiment.

[0013] The preparation method of rabbit anti-bovine lactoferrin serum in this embodiment is as follows: place Freund's complete adjuvant in an agate mortar, and then add 1% to 10% of the mass of Freund's complete adjuvant to the agate mortar, at a concentration of 0.8-1.2 mg / mL bovine lactoferrin standard solution, grind for 3-30 minutes to get the vaccine, then inject 1.2 mg of the vaccine on the hind paw of each rabbit, and inject multiple points under the skin of each rabbit 14 days later 1 mg vaccine, after 20 days, inject 1 mg vaccine at multiple points under the skin of rabbits, and then conduct blood test on rabbits to detect the titer until the titer reaches 1:80000-90000, collect blood from the carotid artery of rabbits, and separate The serum was rab...

specific Embodiment approach 3

[0014] Specific embodiment three: the method for detecting bovine lactoferrin with an enzyme-linked immunosorbent assay kit in this embodiment is carried out according to the following steps: 1. Preparation solution: the detection solution is diluted 1000 times with a sample diluent to obtain a detection diluent, a substrate solution and a mass concentration 0.3% H 2 o 2 Mix according to the ratio of 1:1 to obtain the substrate mixture, and dilute the washing liquid 25 times with deionized water to obtain the washing diluent; 2. Bind the antibody to the enzyme-linked plate: dilute the rabbit anti-bovine lactoferrin serum for 5 times with the sample diluent ~20 times to get rabbit anti-bovine lactoferrin serum dilution, then add 100 μL rabbit anti-bovine lactoferrin serum dilution to each well of the enzyme-linked plate, and then place the enzyme-linked plate at 4°C overnight, that is, the rabbit Anti-bovine lactoferrin serum binds to the enzyme-linked plate; 3. Wash the enzym...

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Abstract

The invention discloses an enzyme-linked immuno kit and a method for detecting bovine lactoferrin by using same, relating to a kit and a method for detecting bovine lactoferrin by using the kit. The invention solves the problems of the existing method for detecting bovine lactoferrin that the detection cost is higher, and the operation is complicated. The enzyme-linked immuno kit comprises an enzyme-linked plate, bovine lactoferrin standard substance, sample diluent, detection solution, substrate solution, cleaning solution and stop solution; the detection method mainly comprises the steps ofpreparing solution, combining an antibody on the enzyme-linked plate, closing the enzyme-linked plate, fixing the standard substance and the sample to be detected on the enzyme-linked plate, detectinglight absorption value by a microplate reader, making a standard curve according to detection results, and then calculating the content of the ovine lactoferrin serous of the sample to be detected. The method adopts a working solution manner, thus having low cost; and the method for detecting bovine lactoferrin by using the enzyme-linked immuno kit is simple in operation.

Description

technical field [0001] The invention relates to a kit and a method for detecting bovine lactoferrin with the kit. Background technique [0002] Bovine lactoferrin is one of the important functional proteins in milk and its products. The consumption of milk and dairy products in my country is increasing year by year. Some people propose that the application of bovine lactoferrin in dairy products, nutritional health products and medicines should be strengthened. Existing bovine lactoferrin detection methods mainly include enzyme-linked immunoassay, high performance liquid chromatography and radioimmunoassay, but these existing bovine lactoferrin detection methods have higher requirements for inspection equipment, resulting in relatively high detection costs. High, operationally complex issues. Contents of the invention [0003] The purpose of the present invention is to provide a kind of enzyme-linked immunosorbent assay kit and the method for detecting bovine lactoferrin ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/543G01N21/31
Inventor 张英华生庆海刘鹏诸晓强迟涛韩秀娥沈阳
Owner 黑龙江省乳品工业技术开发中心
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