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Monoclonal antibody hybridoma cell lines of soybean lipoxygenase isozyme, and antibody and application thereof

A hybridoma cell line, monoclonal antibody technology, applied in biochemical equipment and methods, anti-enzyme immunoglobulin, instruments, etc., can solve problems such as sensitivity and specificity limitations

Active Publication Date: 2013-03-13
INST OF CEREAL & OIL CROPS HEBEI ACAD OF AGRI & FORESTRY SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, since it is determined by the principle of color reaction of lipoxygenase coupled redox indicator, its sensitivity and specificity are limited to a certain extent.

Method used

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  • Monoclonal antibody hybridoma cell lines of soybean lipoxygenase isozyme, and antibody and application thereof
  • Monoclonal antibody hybridoma cell lines of soybean lipoxygenase isozyme, and antibody and application thereof
  • Monoclonal antibody hybridoma cell lines of soybean lipoxygenase isozyme, and antibody and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0072] Embodiment 1: the establishment of the monoclonal antibody hybridoma cell line of anti-soybean lipoxygenase isozyme

[0073] 1.1 Experimental materials:

[0074] Soybean lipoxygenase complete deletion material "Zhongte No. 1" was purchased from the Institute of Crop Science, Chinese Academy of Agricultural Sciences;

[0075] The preservation numbers of the National Resource Bank of Century near-isogenic materials in the United States are: Lox-1: WDD01483, Lox-3: WDD01485;

[0076] Lox-2 enzyme-deficient soybean variety "Wuxing No. 2", approval number: National Examination Bean 2004005;

[0077] BALB / C mice and mouse SP2 / 0 cells were purchased from the Experimental Animal Center of Academy of Military Medical Sciences (Beijing);

[0078] New Zealand white rabbits: purchased from the Experimental Animal Center of Hebei Province;

[0079] 96-well polystyrene microplates were purchased from Xiamen Yunpeng Technology Development Co., Ltd.

[0080] 1.2 Chemical reagents: ...

Embodiment 2

[0105] Example 2: Preparation, detection and application of monoclonal antibodies

[0106] 2.1 Detection antibody preparation

[0107] 2.1.1 Monoclonal antibody preparation and purification

[0108] Preparation of monoclonal antibody ascites: the cell line Lox1-E obtained in step ⑧ in Example 1 1 , Lox2-G 5 , Lox3-H 8 Use culture flasks to expand culture respectively, with 1×10 6 The amount of cells / mouse was inoculated into the peritoneal cavity of BALB / C mice pretreated with paraffin oil to induce ascites. Each cell line was inoculated into 5 mice, and the ascites could be collected after 7 days. Lox1-E 1 , Lox2-G 5 , Lox3-H 8 25ml, 19ml, and 23ml of monoclonal antibody ascites were obtained respectively.

[0109] Ascites titer determination of monoclonal antibody: use indirect ELISA method, Sigma company product Soybean Lox type1-B as detection antigen, determine Lox1-E 1 , Lox2-G 5 , Lox3-H 8 The titers of monoclonal antibodies in ascites were 1:320,000, 1:640,00...

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Abstract

The invention discloses a group of soybean lipoxygenase isozyme resisting monoclonal antibody hybridoma cell lines, wherein the cell lines are respectively a soybean lipoxygenase isozyme Lox1 resisting monoclonal antibody hybridoma cell line with the collection number of CGMCC(China General Microbiological Culture Collection Center) 5121, a soybean lipoxygenase isozyme Lox2 resisting monoclonal antibody hybridoma cell line with the collection number of CGMCC 5122 and a soybean lipoxygenase isozyme Lox3 resisting monoclonal antibody hybridoma cell line with the collection number of CGMCC 5123. The invention further discloses a monoclonal antibody secreted by the hybridoma cell line, and the application of the monoclonal antibody in detection of the soybean lipoxygenase isozyme. The invention provides a method for detecting the soybean lipoxygenase isozyme by using the monoclonal antibody, and a kit is produced according to the method, wherein the kit is suitable for syncrhnously executing qualitative and quantitative determination on three kinds of lipoxidases in the soybean.

Description

technical field [0001] The present invention relates to a kind of monoclonal antibody hybridoma cell line and its monoclonal antibody, particularly relate to the monoclonal antibody hybridoma cell line and its monoclonal antibody of respectively secreting anti-soybean lipoxygenase isozyme Lox1, Lox2 and Lox3, cell The preservation numbers of the strains are: CGMCC 5121, CGMCC 5122 and CGMCC 5123, and also involve the application of the monoclonal antibodies secreted by each of them in the detection of soybean lipoxygenase isozymes by double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA), It belongs to the technical field of biological detection. Background technique [0002] Soybean contains three lipoxygenase (Lox for short) isoenzymes namely Lox1, Lox2 and Lox3, which can specifically catalyze polyunsaturated fatty acids with cis, cis-1, 4-pentadiene structure through molecular addition Oxygen, forming hydroperoxide derivatives with conjugated double bond...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/20C07K16/40G01N33/577G01N33/573
Inventor 张孟臣马志民蒋春志邸锐杨春燕赵双进秦君闫龙刘兵强赵青松唐晓东史晓蕾李亚璞
Owner INST OF CEREAL & OIL CROPS HEBEI ACAD OF AGRI & FORESTRY SCI
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