CN102109516A - Clenbuterol hydrochloride detection kit for polyclonal antibody - Google Patents
Clenbuterol hydrochloride detection kit for polyclonal antibody Download PDFInfo
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- CN102109516A CN102109516A CN2009102006660A CN200910200666A CN102109516A CN 102109516 A CN102109516 A CN 102109516A CN 2009102006660 A CN2009102006660 A CN 2009102006660A CN 200910200666 A CN200910200666 A CN 200910200666A CN 102109516 A CN102109516 A CN 102109516A
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- clenobuterol hydrochloride
- detection kit
- polyclonal antibody
- antibody according
- antiserum
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Abstract
The invention relates to a clenbuterol hydrochloride detection kit for polyclonal antibody. The detection kit consists of a pre-coating reaction plate, a clenbuterol hydrochloride standard substance, an enzyme substrate, an enzyme-labeled secondary antibody, diluent, cleaning solution, stop solution and a working instruction. Compared with the prior art, the clenbuterol hydrochloride detection kit for the polyclonal antibody has the advantages of low manufacturing cost, high specificity and sensitivity, short manufacturing cycle and the like. The rate of cross reaction with ractopamine (rac) and salbutamol (sal) is 0.1 percent, the detection range is 1 ng/mL to 10 mu g/mL, and the kit has high specificity.
Description
Technical field
The present invention relates to a kind of detection kit, especially relate to a kind of clenobuterol hydrochloride detection kit of polyclonal antibody.
Background technology
(Clenbuterol CL), has another name called Celenbuterol to Clenbuterol, is a kind of β
2Receptor stimulating agent can improve the meat production and the price of deed of animal effectively.But, can cause death when serious to internal organs toxic side effects such as the liver of people and animal, kidneys.The Ministry of Agriculture clearly stipulates, a class β such as Clenbuterol
2Receptor stimulating agent is under an embargo and uses as feed addictive.(enzyme-linked immunosorbent assay ELISA) is the common method of present detection by quantitative CL to enzyme linked immunosorbent assay.But the ELISA kit that uses on the market mostly is import, uses the clenobuterol hydrochloride Monoclonal Antibody, costs an arm and a leg, and can not adapt to needed quick, the easy and inexpensive requirement of market surveillance.
Summary of the invention
Purpose of the present invention is exactly to provide in order to overcome the defective that above-mentioned prior art exists that a kind of cost is lower, the clenobuterol hydrochloride detection kit of the polyclonal antibody of high specificity.
Purpose of the present invention can be achieved through the following technical solutions:
A kind of clenobuterol hydrochloride detection kit of polyclonal antibody is characterized in that, this test kit is made up of following: wrap by reaction plate, clenobuterol hydrochloride standard items, zymolyte, ELIAS secondary antibody, dilution, cleansing solution and stop buffer in advance.
Described pre-bag is prepared by following steps by reaction plate:
(1) with isodose Fu Shi Freund's complete adjuvant emulsification clenobuterol hydrochloride-bovine serum albumin(BSA) conjugate, to 6 the week age BALB/C female mice carry out subcutaneous and abdominal cavity multi-point injection immunity, then to the mouse blood sampling of docking, separate and obtain antiserum, survey antiserum titre and specific detection with indirect ELISA, it is standby that qualified antiserum is stored in-20 ℃ of preservations;
(2) with clenobuterol hydrochloride-chicken egg white conjugate coated elisa plate, the control temperature is 4 ℃ spends the night, and washs then, pats dry, and adds the good antiserum 100 μ L of dilution in each hole of coated elisa plate, place 1h, wash, pat dry and promptly make pre-bag for 37 ℃ by reaction plate.
Antiserum titre qualified in the described step (1) is 1 * 10
5More than, and with salbutamol and Ractopamine no cross reaction.
The sero-fast extension rate that dilution is good in the described step (2) is 10000-13000.
The concentration of clenobuterol hydrochloride is 1 μ g/ μ L in the described clenobuterol hydrochloride standard items.
Described zymolyte is that concentration is the o-phenylenediamine solution of 0.5mg/ml.
Described ELIAS secondary antibody is that the horseradish peroxidase mark two of sheep anti mouse is anti-.
Described dilution is that pH is 7.4 phosphate buffer.
Described cleansing solution is for containing 0.05wt%Tween 20, and pH is 7.4 phosphate buffer.
Described stop buffer is the concentrated sulphuric acid of 98wt%.
Compared with prior art, it is lower that the present invention has cost of manufacture, advantages such as high specificity, highly sensitive and fabrication cycle are short, with Ractopamine (Ractopamine, Rac), salbutamol (Salbutamol, Sal) cross reacting rate is 0.1%, and sensing range is 1ng/mL-10 μ g/mL, has specificity preferably.
Embodiment
The present invention is described in detail below in conjunction with specific embodiment.
Embodiment 1
A kind of clenobuterol hydrochloride detection kit of polyclonal antibody, this test kit is made up of following: wrap in advance by reaction plate, the clenobuterol hydrochloride standard items, zymolyte, ELIAS secondary antibody, dilution, cleansing solution, stop buffer and operation instructions, in the kit in the clenobuterol hydrochloride standard items concentration of clenobuterol hydrochloride be 1 μ g/ μ L, zymolyte is the o-phenylenediamine solution of 0.5mg/ml, ELIAS secondary antibody is that the horseradish peroxidase mark two of sheep anti mouse is anti-, dilution is the phosphate buffer of pH=7.4, cleansing solution is for containing the 0.05wt% polysorbas20, the phosphate buffer of pH=7.4, stop buffer are the concentrated sulphuric acid of 98wt%.
Pre-bag is prepared by following steps by reaction plate:
(1) with isodose Fu Shi Freund's complete adjuvant emulsification clenobuterol hydrochloride-bovine serum albumin(BSA) conjugate, to 6 the week age BALB/C female mice carry out subcutaneous and abdominal cavity multi-point injection immunity, then to the mouse blood sampling of docking, separate and obtain antiserum, survey antiserum titre and specific detection with indirect ELISA, it is standby that qualified antiserum is stored in-20 ℃ of preservations, and this qualified antiserum titre is 1 * 10
5More than, and with salbutamol and Ractopamine no cross reaction;
(2) with clenobuterol hydrochloride-chicken egg white conjugate coated elisa plate, the control temperature is 4 ℃ spends the night, and washs then, pats dry, and adds the antiserum 100 μ L of 10000 times of dilutions in each hole of coated elisa plate, place 1h, wash, pat dry and promptly make pre-bag for 37 ℃ by reaction plate.
Embodiment 2
A kind of clenobuterol hydrochloride detection kit of polyclonal antibody, this test kit is made up of following: wrap in advance by reaction plate, the clenobuterol hydrochloride standard items, zymolyte, ELIAS secondary antibody, dilution, cleansing solution, stop buffer and operation instructions, in the kit in the clenobuterol hydrochloride standard items concentration of clenobuterol hydrochloride be 1 μ g/ μ L, zymolyte is the o-phenylenediamine solution of 0.5mg/ml, ELIAS secondary antibody is that the horseradish peroxidase mark two of sheep anti mouse is anti-, dilution is the phosphate buffer of pH=7.4, cleansing solution is for containing the 0.05wt% polysorbas20, the phosphate buffer of pH=7.4, stop buffer are the concentrated sulphuric acid of 98wt%.
Pre-bag is prepared by following steps by reaction plate:
(1) with isodose Fu Shi Freund's complete adjuvant emulsification clenobuterol hydrochloride-bovine serum albumin(BSA) conjugate, to 6 the week age BALB/C female mice carry out subcutaneous and abdominal cavity multi-point injection immunity, then to the mouse blood sampling of docking, separate and obtain antiserum, survey antiserum titre and specific detection with indirect ELISA, it is standby that qualified antiserum is stored in-20 ℃ of preservations, and this qualified antiserum titre is 1 * 10
5More than, and with salbutamol and Ractopamine no cross reaction;
(2) with clenobuterol hydrochloride-chicken egg white conjugate coated elisa plate, the control temperature is 4 ℃ spends the night, and washs then, pats dry, and adds the antiserum 100 μ L of 13000 times of dilutions in each hole of coated elisa plate, place 1h, wash, pat dry and promptly make pre-bag for 37 ℃ by reaction plate.
Claims (10)
1. the clenobuterol hydrochloride detection kit of a polyclonal antibody is characterized in that, this test kit is made up of following: wrap by reaction plate, clenobuterol hydrochloride standard items, zymolyte, ELIAS secondary antibody, dilution, cleansing solution and stop buffer in advance.
2. the clenobuterol hydrochloride detection kit of a kind of polyclonal antibody according to claim 1 is characterized in that, described pre-bag is prepared by following steps by reaction plate:
(1) with isodose Fu Shi Freund's complete adjuvant emulsification clenobuterol hydrochloride-bovine serum albumin(BSA) conjugate, to 6 the week age BALB/C female mice carry out subcutaneous and abdominal cavity multi-point injection immunity, then to the mouse blood sampling of docking, separate and obtain antiserum, survey antiserum titre and specific detection with indirect ELISA, it is standby that qualified antiserum is stored in-20 ℃ of preservations;
(2) with clenobuterol hydrochloride-chicken egg white conjugate coated elisa plate, the control temperature is 4 ℃ spends the night, and washs then, pats dry, and adds the good antiserum 100 μ L of dilution in each hole of coated elisa plate, place 1h, wash, pat dry and promptly make pre-bag for 37 ℃ by reaction plate.
3. the clenobuterol hydrochloride detection kit of a kind of polyclonal antibody according to claim 2 is characterized in that, antiserum titre qualified in the described step (1) is 1 * 10
5More than, and with salbutamol and Ractopamine no cross reaction.
4. the clenobuterol hydrochloride detection kit of a kind of polyclonal antibody according to claim 2 is characterized in that, the sero-fast extension rate that dilution is good in the described step (2) is 10000-13000.
5. the clenobuterol hydrochloride detection kit of a kind of polyclonal antibody according to claim 1 is characterized in that, the concentration of clenobuterol hydrochloride is 1 μ g/ μ L in the described clenobuterol hydrochloride standard items.
6. the clenobuterol hydrochloride detection kit of a kind of polyclonal antibody according to claim 1 is characterized in that, described zymolyte is that concentration is the o-phenylenediamine solution of 0.5mg/ml.
7. the clenobuterol hydrochloride detection kit of a kind of polyclonal antibody according to claim 1 is characterized in that, described ELIAS secondary antibody is that the horseradish peroxidase mark two of sheep anti mouse is anti-.
8. the clenobuterol hydrochloride detection kit of a kind of polyclonal antibody according to claim 1 is characterized in that, described dilution is that pH is 7.4 phosphate buffer.
9. the clenobuterol hydrochloride detection kit of a kind of polyclonal antibody according to claim 1 is characterized in that, described cleansing solution is for containing 0.05wt%Tween 20, and pH is 7.4 phosphate buffer.
10. the clenobuterol hydrochloride detection kit of a kind of polyclonal antibody according to claim 1 is characterized in that, described stop buffer is the concentrated sulphuric acid of 98wt%.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2009102006660A CN102109516A (en) | 2009-12-24 | 2009-12-24 | Clenbuterol hydrochloride detection kit for polyclonal antibody |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2009102006660A CN102109516A (en) | 2009-12-24 | 2009-12-24 | Clenbuterol hydrochloride detection kit for polyclonal antibody |
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|---|---|
| CN102109516A true CN102109516A (en) | 2011-06-29 |
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| Application Number | Title | Priority Date | Filing Date |
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| CN2009102006660A Pending CN102109516A (en) | 2009-12-24 | 2009-12-24 | Clenbuterol hydrochloride detection kit for polyclonal antibody |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102520155A (en) * | 2011-12-13 | 2012-06-27 | 潍坊市康华生物技术有限公司 | Clenbuterol hydrochloride assay kit and its preparation method and use method |
| CN103226146A (en) * | 2012-01-30 | 2013-07-31 | 广州瑞森生物科技有限公司 | Clenbuterol, ractopamine and salbutamol three-in-one euzyme linked immunosorbent assay, special kit and kit detection method |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2003091290A1 (en) * | 2002-04-23 | 2003-11-06 | Boehringer Ingelheim Pharmaceuticals, Inc. | Method for reduction of residual organic solvent in carbomer |
| CN1885038A (en) * | 2006-07-11 | 2006-12-27 | 华南农业大学 | ELISA kit for detecting clenbuterol and detection method thereof, and animal tissue sample preparing method before detection |
| CN101368952A (en) * | 2008-09-24 | 2009-02-18 | 四川大学 | ELISA adsorption analysis method for measuring clenobuterol hydrochloride content in milk, pork liver, chicken liver and animal feed |
| CN101413951A (en) * | 2008-11-27 | 2009-04-22 | 上海交通大学 | Chemiluminescence immune detection reagent kit for detecting Clenbuterol |
-
2009
- 2009-12-24 CN CN2009102006660A patent/CN102109516A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2003091290A1 (en) * | 2002-04-23 | 2003-11-06 | Boehringer Ingelheim Pharmaceuticals, Inc. | Method for reduction of residual organic solvent in carbomer |
| CN1885038A (en) * | 2006-07-11 | 2006-12-27 | 华南农业大学 | ELISA kit for detecting clenbuterol and detection method thereof, and animal tissue sample preparing method before detection |
| CN101368952A (en) * | 2008-09-24 | 2009-02-18 | 四川大学 | ELISA adsorption analysis method for measuring clenobuterol hydrochloride content in milk, pork liver, chicken liver and animal feed |
| CN101413951A (en) * | 2008-11-27 | 2009-04-22 | 上海交通大学 | Chemiluminescence immune detection reagent kit for detecting Clenbuterol |
Non-Patent Citations (2)
| Title |
|---|
| 陈存社等: "ELISA检测盐酸克伦特罗残留的方法学评价", 《食品科学》 * |
| 陈存社等: "盐酸克伦特罗残留酶联免疫吸附(ELISA)检测方法的研究", 《食品与发酵工业》 * |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102520155A (en) * | 2011-12-13 | 2012-06-27 | 潍坊市康华生物技术有限公司 | Clenbuterol hydrochloride assay kit and its preparation method and use method |
| CN102520155B (en) * | 2011-12-13 | 2013-11-06 | 潍坊市康华生物技术有限公司 | Clenbuterol hydrochloride assay kit and its preparation method and use method |
| CN103226146A (en) * | 2012-01-30 | 2013-07-31 | 广州瑞森生物科技有限公司 | Clenbuterol, ractopamine and salbutamol three-in-one euzyme linked immunosorbent assay, special kit and kit detection method |
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Application publication date: 20110629 |