CN106353313B - A kind of remaining quick detection kit of melamine in dairy products and application method - Google Patents
A kind of remaining quick detection kit of melamine in dairy products and application method Download PDFInfo
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- CN106353313B CN106353313B CN201610983887.XA CN201610983887A CN106353313B CN 106353313 B CN106353313 B CN 106353313B CN 201610983887 A CN201610983887 A CN 201610983887A CN 106353313 B CN106353313 B CN 106353313B
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- agent
- melamine
- dairy products
- buffer
- ultrafiltration
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/75—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated
- G01N21/77—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator
- G01N21/78—Systems in which material is subjected to a chemical reaction, the progress or the result of the reaction being investigated by observing the effect on a chemical indicator producing a change of colour
Abstract
It include inorganic agent, color developing agent, extractant, dropper, buffer, reducing agent and enzyme agent the invention discloses a kind of remaining quick detection kit of melamine in dairy products and application method, the inorganic agent is trichloroacetic acid and phenol, the color developing agent is the mixture of potassium mercuric iodide, phosphomolybdic acid and thiocarbamide, the extractant is sodium acetate solution, the buffer is Gly-HCl buffer, the reducing agent is dithiothreitol (DTT) and beta -mercaptoethanol, and the enzyme agent is plant esterase.The present invention is at low cost, production is simple, detection sensitivity is high, its remolding sensitivity prior art is higher by decades of times and thousands of times, and the detection range of linearity is significantly broadening, it is easy to use, it is simple and quick, complicated reagent preparation process is not needed in testing, and layman can be readily accomplished detection according to kit specification.
Description
Technical field
The present invention relates to a kind of kit, the remaining quick detection kit of specifically a kind of melamine in dairy products and
Application method.
Background technique
Melamine is commonly called as melamine etc., it is a kind of synthesis organic nitrogen-containing heterocyclic compound, belongs to a kind of industrial chemicals,
It is widely applied in the fields such as textile industry, paper-making industry and packaging material.Melamine can be from the links such as ring mirror and packaging material for food
In enter in food, but in general, its content is lower.A kind of white crystalline powder of melamine, nitrogen content are very high
(66%), protein content can be significantly improved, people are referred to as " extract of protein ".In addition its simple production process, cost are very low.Cause
The illegal manufacturer in this part is driven by interests, and melamine is added in the food such as milk, to significantly improve the egg detected
White matter content.But the melamine artificially added is far beyond the dosis tolerata of people, into body after easily formed it is not soluble in water
Macromolecular complex deposition and form calculus, damage kidney, serious person can cause renal failure, and research also demonstrates, melamine
A large amount of intakes of amine also will cause the damage of reproductive system, while also result in the generation of the serious diseases such as bladder cancer, seriously threaten
The health and life security of people.
The detection method of melamine in currently available technology is more, including near-infrared, middle infrared technique, enzyme-linked exempts from
Epidemic disease, but these methods exist to instrument requirements height, the disadvantage that testing cost is high, false positive rate is high.Therefore, in order to accurately and effectively
The melamine in food is detected, guarantees food safety and people's health, urgent need seeks a kind of detection rapidly, high sensitivity
Melamine detection method.
Summary of the invention
That the purpose of the present invention is to provide a kind of structures is simple, melamine in dairy products easy to use is remaining quickly
Detection kit and application method, to solve the problems mentioned in the above background technology.
To achieve the above object, the invention provides the following technical scheme:
A kind of remaining quick detection kit of melamine in dairy products, including inorganic agent, color developing agent, extractant, drop
Pipe, buffer, reducing agent and enzyme agent, the inorganic agent are trichloroacetic acid and phenol, and the color developing agent is potassium mercuric iodide, phosphomolybdic acid
With the mixture of thiocarbamide, the extractant is sodium acetate solution, and the buffer is Gly-HCl buffer, and the reducing agent is
Dithiothreitol (DTT) and beta -mercaptoethanol, the enzyme agent are plant esterase.
A kind of application method of the remaining quick detection kit of melamine in dairy products, the specific steps are as follows:
(1) it firstly, dairy products are sealed off on aseptic operating platform, is placed in mortar after drawing 10-20ml, addition mentions
It takes agent and liquid nitrogen quickly to be ground, is then added drop-wise to 96 hole elisa Plates using the solution that dropper is drawn after 50-100 μ L grinding
In, it is then placed in -80 DEG C of refrigerators of ultralow temperature and refrigerate for use；
(2) then, sodium tartrate, bovine serum albumin, lanthanum chloride, oxalic acid are added in deoxynucleotide sodium solution, are filled
Divide and stir evenly, placement is protected from light place's cryo-conservation；
(3) then, 96 stand-by hole elisa Plates will be refrigerated in above-mentioned steps (1) to take out, buffer is added after defrosting and goes back
Former agent, wherein 4000-6000 times of dilution agent of buffering, wherein standing 15min at 600-800 times of dilution agent, 4 DEG C of reduction；
(4) then, will in above-mentioned steps (2) uniformly after deoxynucleotide sodium solution carry out ultrafiltration, after ultrafiltration stand to
With；
(5) then, addition inorganic agent, enzyme agent and color developing agent in 96 hole elisa Plates after above-mentioned steps (3) being stood, 37 DEG C
Then the deoxynucleotide sodium solution after ultrafiltration is added in lower incubation 20min, after mixing well, observe color change；
(6) finally, the color after variation is compared using standard color comparison card.
As a further solution of the present invention: the mass percent concentration of deoxynucleotide sodium solution is in step (2)
4.6%。
As a further solution of the present invention: bovine serum albumin is from Zhejiang day Hangzhoupro biotechnology share in step (2)
Co., Ltd.
As a further solution of the present invention: ultrafiltration is specially to use interception for 200-300KDa in step (4)
Ultrafiltration membrane carries out ultrafiltration.
As a further solution of the present invention: enzyme agent is using preceding needing to be preheating to 37 DEG C in step (5).
Compared with prior art, the beneficial effects of the present invention are:
The present invention is at low cost, production is simple, and detection sensitivity is high, and the remolding sensitivity prior art is higher by decades of times and thousands of
Times, and the detection range of linearity is significantly broadening, and it is easy to use, it is simple and quick, complicated reagent preparation is not needed in testing
Process, layman can be readily accomplished detection according to kit specification.
Specific embodiment
The technical solution of the patent is explained in further detail With reference to embodiment.
Embodiment 1
A kind of remaining quick detection kit of melamine in dairy products, including inorganic agent, color developing agent, extractant, drop
Pipe, buffer, reducing agent and enzyme agent, the inorganic agent are trichloroacetic acid and phenol, and the color developing agent is potassium mercuric iodide, phosphomolybdic acid
With the mixture of thiocarbamide, the extractant is sodium acetate solution, and the buffer is Gly-HCl buffer, and the reducing agent is
Dithiothreitol (DTT) and beta -mercaptoethanol, the enzyme agent are plant esterase.
A kind of application method of the remaining quick detection kit of melamine in dairy products, the specific steps are as follows:
(1) it firstly, dairy products are sealed off on aseptic operating platform, is placed in mortar after drawing 10ml, is added and extracts
Agent and liquid nitrogen are quickly ground, and are then added drop-wise in 96 hole elisa Plates using the solution that dropper is drawn after 50 μ L grinding, then
It is placed in -80 DEG C of refrigerators of ultralow temperature and refrigerate for use；
(2) then, sodium tartrate, bovine serum albumin, lanthanum chloride, oxalic acid are added in deoxynucleotide sodium solution, are filled
Divide and stir evenly, placement is protected from light place's cryo-conservation；
(3) then, 96 stand-by hole elisa Plates will be refrigerated in above-mentioned steps (1) to take out, buffer is added after defrosting and goes back
Former agent, wherein 4000 times of dilution agent of buffering, wherein standing 15min at 600 times of dilution agent, 4 DEG C of reduction；
(4) then, will in above-mentioned steps (2) uniformly after deoxynucleotide sodium solution carry out ultrafiltration, after ultrafiltration stand to
With；
(5) then, addition inorganic agent, enzyme agent and color developing agent in 96 hole elisa Plates after above-mentioned steps (3) being stood, 37 DEG C
Then the deoxynucleotide sodium solution after ultrafiltration is added in lower incubation 20min, after mixing well, observe color change；
(6) finally, the color after variation is compared using standard color comparison card.
Wherein:
The mass percent concentration of deoxynucleotide sodium solution is 4.6% in step (2).
Bovine serum albumin derives from the biotech inc Zhejiang Tian Hang in step (2).
Ultrafiltration is specially that interception is used to carry out ultrafiltration for 200-300KDa ultrafiltration membrane in step (4).
Enzyme agent is using preceding needing to be preheating to 37 DEG C in step (5).
Embodiment 2
A kind of remaining quick detection kit of melamine in dairy products, including inorganic agent, color developing agent, extractant, drop
Pipe, buffer, reducing agent and enzyme agent, the inorganic agent are trichloroacetic acid and phenol, and the color developing agent is potassium mercuric iodide, phosphomolybdic acid
With the mixture of thiocarbamide, the extractant is sodium acetate solution, and the buffer is Gly-HCl buffer, and the reducing agent is
Dithiothreitol (DTT) and beta -mercaptoethanol, the enzyme agent are plant esterase.
A kind of application method of the remaining quick detection kit of melamine in dairy products, the specific steps are as follows:
(1) it firstly, dairy products are sealed off on aseptic operating platform, is placed in mortar after drawing 15ml, is added and extracts
Agent and liquid nitrogen are quickly ground, and are then added drop-wise in 96 hole elisa Plates using the solution that dropper is drawn after 80 μ L grinding, then
It is placed in -80 DEG C of refrigerators of ultralow temperature and refrigerate for use；
(2) then, sodium tartrate, bovine serum albumin, lanthanum chloride, oxalic acid are added in deoxynucleotide sodium solution, are filled
Divide and stir evenly, placement is protected from light place's cryo-conservation；
(3) then, 96 stand-by hole elisa Plates will be refrigerated in above-mentioned steps (1) to take out, buffer is added after defrosting and goes back
Former agent, wherein 5000 times of dilution agent of buffering, wherein standing 15min at 700 times of dilution agent, 4 DEG C of reduction；
(4) then, will in above-mentioned steps (2) uniformly after deoxynucleotide sodium solution carry out ultrafiltration, after ultrafiltration stand to
With；
(5) then, addition inorganic agent, enzyme agent and color developing agent in 96 hole elisa Plates after above-mentioned steps (3) being stood, 37 DEG C
Then the deoxynucleotide sodium solution after ultrafiltration is added in lower incubation 20min, after mixing well, observe color change；
(6) finally, the color after variation is compared using standard color comparison card.
Wherein:
The mass percent concentration of deoxynucleotide sodium solution is 4.6% in step (2).
Bovine serum albumin derives from the biotech inc Zhejiang Tian Hang in step (2).
Ultrafiltration is specially that interception is used to carry out ultrafiltration for 200-300KDa ultrafiltration membrane in step (4).
Enzyme agent is using preceding needing to be preheating to 37 DEG C in step (5).
Embodiment 3
A kind of remaining quick detection kit of melamine in dairy products, including inorganic agent, color developing agent, extractant, drop
Pipe, buffer, reducing agent and enzyme agent, the inorganic agent are trichloroacetic acid and phenol, and the color developing agent is potassium mercuric iodide, phosphomolybdic acid
With the mixture of thiocarbamide, the extractant is sodium acetate solution, and the buffer is Gly-HCl buffer, and the reducing agent is
Dithiothreitol (DTT) and beta -mercaptoethanol, the enzyme agent are plant esterase.
A kind of application method of the remaining quick detection kit of melamine in dairy products, the specific steps are as follows:
(1) it firstly, dairy products are sealed off on aseptic operating platform, is placed in mortar after drawing 20ml, is added and extracts
Agent and liquid nitrogen are quickly ground, and are then added drop-wise in 96 hole elisa Plates using the solution that dropper is drawn after 100 μ L grinding, then
It is placed in -80 DEG C of refrigerators of ultralow temperature and refrigerate for use；
(2) then, sodium tartrate, bovine serum albumin, lanthanum chloride, oxalic acid are added in deoxynucleotide sodium solution, are filled
Divide and stir evenly, placement is protected from light place's cryo-conservation；
(3) then, 96 stand-by hole elisa Plates will be refrigerated in above-mentioned steps (1) to take out, buffer is added after defrosting and goes back
Former agent, wherein 6000 times of dilution agent of buffering, wherein standing 15min at 800 times of dilution agent, 4 DEG C of reduction；
(4) then, will in above-mentioned steps (2) uniformly after deoxynucleotide sodium solution carry out ultrafiltration, after ultrafiltration stand to
With；
(5) then, addition inorganic agent, enzyme agent and color developing agent in 96 hole elisa Plates after above-mentioned steps (3) being stood, 37 DEG C
Then the deoxynucleotide sodium solution after ultrafiltration is added in lower incubation 20min, after mixing well, observe color change；
(6) finally, the color after variation is compared using standard color comparison card.
Wherein:
The mass percent concentration of deoxynucleotide sodium solution is 4.6% in step (2).
Bovine serum albumin derives from the biotech inc Zhejiang Tian Hang in step (2).
Ultrafiltration is specially that interception is used to carry out ultrafiltration for 200-300KDa ultrafiltration membrane in step (4).
Enzyme agent is using preceding needing to be preheating to 37 DEG C in step (5).
The present invention is at low cost, production is simple, and detection sensitivity is high, and the remolding sensitivity prior art is higher by decades of times and thousands of
Times, and the detection range of linearity is significantly broadening, and it is easy to use, it is simple and quick, complicated reagent preparation is not needed in testing
Process, layman can be readily accomplished detection according to kit specification.
The preferred embodiment of the patent is described in detail above, but this patent is not limited to above-mentioned embodiment party
Formula within the knowledge of one of ordinary skill in the art can also be under the premise of not departing from this patent objective
Various changes can be made.
Claims (5)
1. a kind of remaining quick detection kit of melamine in dairy products, which is characterized in that including inorganic agent, color developing agent,
Extractant, dropper, buffer, reducing agent and enzyme agent, the inorganic agent are trichloroacetic acid and phenol, and the color developing agent is mercuric iodixde
The mixture of potassium, phosphomolybdic acid and thiocarbamide, the extractant are sodium acetate solution, and the buffer is Gly-HCl buffer, described
Reducing agent is dithiothreitol (DTT) and beta -mercaptoethanol, and the enzyme agent is plant esterase.
2. a kind of application method of the remaining quick detection kit of melamine in dairy products as described in claim 1,
It is characterized in that, the specific steps are as follows:
(1) it firstly, dairy products are sealed off on aseptic operating platform, is placed in mortar after drawing 10-20ml, extractant is added
It is quickly ground, is then added drop-wise in 96 hole elisa Plates using the solution that dropper is drawn after 50-100 μ L grinding, so with liquid nitrogen
After be placed in -80 DEG C of refrigerators of ultralow temperature refrigerate it is stand-by；
(2) then, sodium tartrate, bovine serum albumin, lanthanum chloride, oxalic acid are added in deoxynucleotide sodium solution, are sufficiently stirred
It mixes uniformly, placement is protected from light place's cryo-conservation；
(3) then, 96 stand-by hole elisa Plates of refrigeration in above-mentioned steps (1) are taken out, buffer and reducing agent are added after defrosting,
4000-6000 times of dilution agent is wherein buffered, wherein standing 15min at 600-800 times of dilution agent, 4 DEG C of reduction；
(4) then, the deoxynucleotide sodium solution after will be uniform in above-mentioned steps (2) carries out ultrafiltration, set aside for use after ultrafiltration；
(5) then, inorganic agent, enzyme agent and color developing agent are added in 96 hole elisa Plates after above-mentioned steps (3) being stood, is incubated at 37 DEG C
20min is educated, the deoxynucleotide sodium solution after ultrafiltration is then added, after mixing well, observes color change；
(6) finally, the color after variation is compared using standard color comparison card.
3. the application method of the remaining quick detection kit of melamine in dairy products according to claim 2, special
Sign is that the mass percent concentration of deoxynucleotide sodium solution is 4.6% in step (2).
4. the application method of the remaining quick detection kit of melamine in dairy products according to claim 2, special
Sign is that ultrafiltration is specially that interception is used to carry out ultrafiltration for 200-300KDa ultrafiltration membrane in step (4).
5. the application method of the remaining quick detection kit of melamine in dairy products according to claim 2, special
Sign is, enzyme agent is using preceding needing to be preheating to 37 DEG C in step (5).
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| CN201610983887.XA CN106353313B (en) | 2016-11-09 | 2016-11-09 | A kind of remaining quick detection kit of melamine in dairy products and application method |
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| CN101881759A (en) * | 2009-05-05 | 2010-11-10 | 天津生机集团股份有限公司 | Tripolycyanamide rapid detection kit and application thereof |
| CN102519951A (en) * | 2011-12-06 | 2012-06-27 | 江南大学 | Color reagent made from colorimetry nanometer material and used for detecting melamine and application of color reagent |
| CN102954959A (en) * | 2011-08-29 | 2013-03-06 | 中国科学院上海有机化学研究所 | Melamine colorimetric detection method based on enzymatic amino hydrolysis reaction |
| CN102998305A (en) * | 2011-09-19 | 2013-03-27 | 青岛大学 | Method for detecting melamine in raw milk by utilizing chromogenic reaction |
| TWI412494B (en) * | 2009-12-31 | 2013-10-21 | Ind Tech Res Inst | Detection kit of melamine and detection method thereof |
| US8637269B2 (en) * | 2009-07-10 | 2014-01-28 | Bioo Scientific Corporation | Method for the detection of melamine |
| CN103645275A (en) * | 2013-12-03 | 2014-03-19 | 中山鼎晟生物科技有限公司 | Melamine detection reagent and detection method |
| CN104459009A (en) * | 2014-12-09 | 2015-03-25 | 苏州东辰林达检测技术有限公司 | Quick detection kit for melamine and using method thereof |
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2016
- 2016-11-09 CN CN201610983887.XA patent/CN106353313B/en active Active
Patent Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101881759A (en) * | 2009-05-05 | 2010-11-10 | 天津生机集团股份有限公司 | Tripolycyanamide rapid detection kit and application thereof |
| US8637269B2 (en) * | 2009-07-10 | 2014-01-28 | Bioo Scientific Corporation | Method for the detection of melamine |
| TWI412494B (en) * | 2009-12-31 | 2013-10-21 | Ind Tech Res Inst | Detection kit of melamine and detection method thereof |
| CN102954959A (en) * | 2011-08-29 | 2013-03-06 | 中国科学院上海有机化学研究所 | Melamine colorimetric detection method based on enzymatic amino hydrolysis reaction |
| CN102998305A (en) * | 2011-09-19 | 2013-03-27 | 青岛大学 | Method for detecting melamine in raw milk by utilizing chromogenic reaction |
| CN102519951A (en) * | 2011-12-06 | 2012-06-27 | 江南大学 | Color reagent made from colorimetry nanometer material and used for detecting melamine and application of color reagent |
| CN103645275A (en) * | 2013-12-03 | 2014-03-19 | 中山鼎晟生物科技有限公司 | Melamine detection reagent and detection method |
| CN104459009A (en) * | 2014-12-09 | 2015-03-25 | 苏州东辰林达检测技术有限公司 | Quick detection kit for melamine and using method thereof |
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| CN106353313A (en) | 2017-01-25 |
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Effective date of registration: 20190605 Address after: 530000 High-tech Zone, Nanning City, Guangxi Zhuang Autonomous Region, No. 1 High-tech Second Road, Science Park, Guangxi University Patentee after: Agricultural University Food Factory of Guangxi University Address before: 214070 Creative Park No. 100 Dicui Road, Wuxi City, Jiangsu Province, Phase III A Building 303 Patentee before: BEOSEN (JIANGSU) FOOD SAFETY TECHNOLOGY Co.,Ltd. |
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Effective date of registration: 20220729 Address after: 543003 building a-5, Donghua industrial community, Wuzhou Industrial Park, Guangxi Zhuang Autonomous Region Patentee after: Wuzhou Zhongjin Technology Co.,Ltd. Address before: 530000 High-tech Zone, Nanning City, Guangxi Zhuang Autonomous Region, No. 1 High-tech Second Road, Science Park, Guangxi University Patentee before: Agricultural University Food Factory of Guangxi University |
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