CN104311669A - Hybridoma cell strain FQ-E7, imidacloprid-resistant monoclonal antibody produced by hybridoma cell strain FQ-E7 and use of imidacloprid-resistant monoclonal antibody - Google Patents
Hybridoma cell strain FQ-E7, imidacloprid-resistant monoclonal antibody produced by hybridoma cell strain FQ-E7 and use of imidacloprid-resistant monoclonal antibody Download PDFInfo
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- CN104311669A CN104311669A CN201410497755.7A CN201410497755A CN104311669A CN 104311669 A CN104311669 A CN 104311669A CN 201410497755 A CN201410497755 A CN 201410497755A CN 104311669 A CN104311669 A CN 104311669A
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- imidacloprid
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Abstract
The invention relates to hybridoma cell strain FQ-E7, an imidacloprid-resistant monoclonal antibody produced by the hybridoma cell strain FQ-E7 and a use of the imidacloprid-resistant monoclonal antibody. The hybridoma cell strain FQ-E7 is preserved in the China center for type culture collection (CCTCC) and has preservation number of CCTCC NO.C2014107. The imidacloprid-resistant monoclonal antibody secreted by the hybridoma cell strain FQ-E7 can recognize imidacloprid and has high sensitivity and good singularity. An immune-gold localization test strip utilizing the imidacloprid-resistant monoclonal antibody can realize detection of imidacloprid in water, soil and cabbage and has the lowest imidacloprid content detection limit of 2ng/mL.
Description
Technical field
The present invention relates to hybridoma cell strain FQ-E7, the monoclonal antibody of the anti-Provado of its secretion and application.
Background technology
Provado belongs to novel nicotinamide insecticide, moderate toxicity, is made insect dead by the nAChR ligands acted in insect body.The Agricultural pests such as Provado can effectively be prevented eliminating aphis, plant hopper, leafhopper, are widely used in agriculture production.But due to the unreasonable and nonstandard use of agricultural chemicals, while control Agricultural pests, serious threat is brought to environmental health, HUMAN HEALTH and some non-target organisms.Therefore, the method for quick setting up the Determination of Imidacloprid Residue in environment seems more and more important.
At present, the method such as residue detection many employings tlc, vapor-phase chromatography, high performance liquid chromatography of agricultural chemicals.The detectability of these methods is low, highly sensitive, existing relevant technical bid will definitely be for reference, but owing to needing expensive instrument, special operator, and sample pre-treatments is complicated, cost is high, the time is long, therefore can not meet fast and convenient Site Detection requirement better.
Immunologic detection method has fast, cheap, easy, special advantage, can be portable and carry out field monitoring.Overcome the shortcoming of traditional detection method.Set up immunologic detection method, particularly for the immunologic detection method of pesticide molecule, filter out efficient monoclonal antibody the most key.The present invention is the immunologic detection method realizing Provado in environment and agricultural-food, chemosynthesis is for the multiple haptens of Provado, be combined into immunogen with carrier proteins or wrap by source, and filtering out the hybridoma cell strain FQ-E7 of the efficient monoclonal antibody can secreted for Provado.And utilize the monoclonal antibody secreted by this hybridoma cell strain to establish immune-gold labeled ELISA test strip method for Provado in environment and agricultural-food.
Summary of the invention
The object of this invention is to provide hybridoma cell strain FQ-E7, the monoclonal antibody of the anti-Provado of its secretion and application.
Object of the present invention realizes by following technical scheme:
1. hybridoma cell strain FQ-E7 provided by the invention, be preserved in China typical culture collection center (CCTCC) on June 12nd, 2014, preservation address is: China, Wuhan, Wuhan University; Deposit number is CCTCC NO.C2014107.
2. anti-Provado monoclonal antibody, the hybridoma cell strain FQ-E7 that it is CCTCC NO.C2014107 by deposit number secretes and produces.
3. above-mentioned hybridoma cell strain FQ-E7 screens by the following method to obtain:
The synthesis of 3.1 artificial antigens
By chemosynthesis means imidacloprid synthesis haptens: 1-[6-(2-carboxyethylthio-3-pyridine) methyl]-N-nitro-2-tetrahydroglyoxaline imines (H1) and 1-(6-chloro-3-pyridylmethyl)-3-carboxylic propyl group-N-nitro-2-tetrahydroglyoxaline imines (H2).Active ester method is utilized to have the hapten conjugation of C-terminal to (BSA or OVA) on macro-molecular protein.Wherein H1-BSA is as immunogen, and H2-OVA is as wrapping by source.
The screening of 3.2 hybridoma cell strains
With the female BAl BIc/C mouse of the every antigen amount immunity 6-8 health in age in week only of 100 μ g.Each interval time is 2-3 week, after last booster immunization 3 days, takes out mouse boosting cell under aseptic condition, by mouse boosting cell and myeloma cell with 5: 1 cell quantity merge by the method that PEG mediates.Merge and be placed on 37 DEG C, cultivate in 5%CO2 incubator.When culturing cell grows to culture hole 1/3-1/2, with indirect competitive ELISA method filter out height of tiring, inhibiting rate is high, Growth of Cells is vigorous culture hole carries out subclone.After 3-5 subclone, enlarged culturing, sets up hybridoma cell strain.
The preparation of 3.3 monoclonal antibodies
The hybridoma cell strain FQ-E7 of acquisition is injected in advance with the BAlB/C mouse of aseptic paraffin oil process, collects the ascites of this mouse, utilize caprylic acid-ammonium to carry out purification process to the ascites of collecting and obtain anti-Provado monoclonal antibody.
4. semidefinite quantizes the triple channel immuno-chromatographic test paper strip detecting Provado, comprise cardboard, the one side of cardboard once pastes absorbent pad, detecting pad, gold mark pad, sample pad from top to bottom, the overlapping connection in junction of adjacent each pad, described detecting pad take nitrocellulose filter as base wad, and nitrocellulose filter arranges horizontal nature controlling line, detection line I, detection line II and detection line III from top to bottom.Described nature controlling line is coated with rabbit against murine polyclonal antibody, and described detection line I, II and III are coated with Provado haptens-chicken ovalbumin respectively; Described gold mark pad transverse jet scribbles the anti-Provado monoclonal antibody of nano gold mark, and described anti-Provado monoclonal antibody is that the hybridoma cell strain FQ-E7 being CCTCC NO.C2014107 by deposit number secretes generation.
Beneficial effect
(1) Antibody stability of hybridoma cell strain provided by the invention and secretion thereof is high: Cryopreservation of Hybridoma Cells at least keeps 5 years in liquid nitrogen, and the stability of antibody also can be relatively good, and antibody-20 DEG C of refrigerators of purifying at least can deposit 2 years.
(2) monoclonal antibody provided by the invention is highly sensitive: anti-Provado monoclonal antibody provided by the invention can reach 2ng/mL to the lowest detectable limit detecting Provado in solution.
Claims (2)
1., for detecting a monoclonal antibody for Provado, the hybridoma cell strain FQ-E7 being CCTCC NO:C2014107 by deposit number produces.
2. produce the hybridoma cell strain FQ-E7 of the monoclonal antibody for detecting Provado according to claim 1, be preserved in China typical culture collection center on June 12nd, 2014, deposit number is CCTCC NO:C2014107.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410497755.7A CN104311669A (en) | 2014-09-24 | 2014-09-24 | Hybridoma cell strain FQ-E7, imidacloprid-resistant monoclonal antibody produced by hybridoma cell strain FQ-E7 and use of imidacloprid-resistant monoclonal antibody |
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| Application Number | Priority Date | Filing Date | Title |
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| CN201410497755.7A CN104311669A (en) | 2014-09-24 | 2014-09-24 | Hybridoma cell strain FQ-E7, imidacloprid-resistant monoclonal antibody produced by hybridoma cell strain FQ-E7 and use of imidacloprid-resistant monoclonal antibody |
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| CN104311669A true CN104311669A (en) | 2015-01-28 |
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| CN201410497755.7A Pending CN104311669A (en) | 2014-09-24 | 2014-09-24 | Hybridoma cell strain FQ-E7, imidacloprid-resistant monoclonal antibody produced by hybridoma cell strain FQ-E7 and use of imidacloprid-resistant monoclonal antibody |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104974256A (en) * | 2015-04-10 | 2015-10-14 | 南京农业大学 | Anti-thiacloprid monoclonal antibody and application thereof |
| CN105137081A (en) * | 2015-04-28 | 2015-12-09 | 南京农业大学 | Monoclonal antibody-based imidacloprid detection test paper strip |
| CN105181949A (en) * | 2015-09-22 | 2015-12-23 | 浙江大学 | Rapid test method for 3 kinds of pesticide residue such as imidacloprid in tea and test paper strip used in rapid test method |
| CN105675874A (en) * | 2016-01-11 | 2016-06-15 | 北京勤邦生物技术有限公司 | Colloidal gold test strip for detection of imidacloprid and application thereof |
| CN105784994A (en) * | 2016-03-07 | 2016-07-20 | 中国烟草总公司贵州省公司 | Test paper for detecting imidacloprid residues in crops and application and preparing method thereof |
| CN114591916A (en) * | 2022-03-23 | 2022-06-07 | 扬州大学 | Cell strain secreting anti-imidacloprid monoclonal antibody, monoclonal antibody thereof and application of monoclonal antibody |
| CN115246818A (en) * | 2022-05-25 | 2022-10-28 | 中国医学科学院药用植物研究所 | Hapten, artificial antigen and antibody for detecting imidacloprid as well as preparation methods and applications thereof |
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| KR100696615B1 (en) * | 2005-10-06 | 2007-03-19 | 충북대학교 산학협력단 | Analysing method of insecticide imidacloprid and measurement control system for the same |
| CN101880325A (en) * | 2010-06-22 | 2010-11-10 | 南京农业大学 | Monoclonal antibody for detecting imidacloprid pesticide residue |
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2014
- 2014-09-24 CN CN201410497755.7A patent/CN104311669A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| KR100696615B1 (en) * | 2005-10-06 | 2007-03-19 | 충북대학교 산학협력단 | Analysing method of insecticide imidacloprid and measurement control system for the same |
| CN101880325A (en) * | 2010-06-22 | 2010-11-10 | 南京农业大学 | Monoclonal antibody for detecting imidacloprid pesticide residue |
Cited By (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104974256A (en) * | 2015-04-10 | 2015-10-14 | 南京农业大学 | Anti-thiacloprid monoclonal antibody and application thereof |
| CN104974256B (en) * | 2015-04-10 | 2018-06-05 | 南京农业大学 | A kind of anti-thiacloprid monoclonal antibody and application thereof |
| CN105137081A (en) * | 2015-04-28 | 2015-12-09 | 南京农业大学 | Monoclonal antibody-based imidacloprid detection test paper strip |
| CN105181949A (en) * | 2015-09-22 | 2015-12-23 | 浙江大学 | Rapid test method for 3 kinds of pesticide residue such as imidacloprid in tea and test paper strip used in rapid test method |
| CN105181949B (en) * | 2015-09-22 | 2017-09-15 | 浙江大学 | Rapid test method for 3 kinds of pesticide residue such as imidacloprid in tea and test paper strip used in rapid test method |
| CN105675874A (en) * | 2016-01-11 | 2016-06-15 | 北京勤邦生物技术有限公司 | Colloidal gold test strip for detection of imidacloprid and application thereof |
| CN105784994A (en) * | 2016-03-07 | 2016-07-20 | 中国烟草总公司贵州省公司 | Test paper for detecting imidacloprid residues in crops and application and preparing method thereof |
| CN114591916A (en) * | 2022-03-23 | 2022-06-07 | 扬州大学 | Cell strain secreting anti-imidacloprid monoclonal antibody, monoclonal antibody thereof and application of monoclonal antibody |
| CN114591916B (en) * | 2022-03-23 | 2023-05-16 | 扬州大学 | Cell strain secreting anti-imidacloprid monoclonal antibody, monoclonal antibody thereof and application thereof |
| CN115246818A (en) * | 2022-05-25 | 2022-10-28 | 中国医学科学院药用植物研究所 | Hapten, artificial antigen and antibody for detecting imidacloprid as well as preparation methods and applications thereof |
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Application publication date: 20150128 |