CN104007104A - Kit for rapidly detecting organophosphorus pesticide residues and application method thereof - Google Patents
Kit for rapidly detecting organophosphorus pesticide residues and application method thereof Download PDFInfo
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- CN104007104A CN104007104A CN201410219923.6A CN201410219923A CN104007104A CN 104007104 A CN104007104 A CN 104007104A CN 201410219923 A CN201410219923 A CN 201410219923A CN 104007104 A CN104007104 A CN 104007104A
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Abstract
The invention relates to a kit for rapidly detecting organophosphorus pesticide residues and an application method thereof. The kit comprises a C enzyme bottle, a T enzyme bottle, a C substrate reaction cup and a T substrate reaction cup, wherein the C enzyme bottle and the T enzyme bottle are sealed by using enzyme bottle sealing covers and are filled with an enzyme lyophilized three-enzyme mixing agent; the C substrate reaction cup and the T substrate reaction cup are provided with lyophilization and water-absorption cushions containing acetyl choline, luminol and dimethylbenzidine. The invention further relates to a method for applying the kit to rapidly detecting the organophosphorus pesticide residues. The kit takes acetylcholin esterase, choline oxidase and peroxidase as a catalysis system; a product obtained by hydrolyzing and oxidizing acetylcholine chloride and luminol have a chemiluminescence effect. A method for flexibly testing organophosphorus pesticides in agricultural products is established. The kit has the advantages of high detection speed, simple operation process, good stability and repeatability, no need of expensive instruments and the like, and is very suitable for carrying out field monitoring on the organophosphorus pesticides in farm markets.
Description
Technical field
The present invention relates to the method for organophosphorus pesticide in a kind of fast detecting food, specifically relate to a kind of kit and using method thereof of fast detecting organophosphorus pesticide.
Background technology
Residues of pesticides in agricultural product, food and medicine are still one of important latency of danger side of body human body health.In Chinese agricultural production process, in agricultural chemicals year use amount, still occupy first place in the world, and the use amount of organophosphorus pesticide accounts for the more than 70% of whole pesticide dosage.According to document announcement in recent years, vegetables, fruit organophosphorus pesticide disqualification rate that in the metropolitan market of farm produce of Chinese Provincial, the first quarter sells account for more than 14%, and the persticide residue that especially comes from greenhouse gardening vegetables is higher.In agricultural chemicals sampling monitoring, find, the recall rate of acephatemet, metrifonate, flolimat, DDVP is higher.The method of current detection organophosphorus pesticide mainly contains: 1. spectroscopic methodology, mainly comprises spectrophotometric method, chemoluminescence method, infrared and fluorescent spectrometry; 2. red, orange, green, blue, yellow (ROGBY), mainly comprises thin layer chromatography, vapor-phase chromatography, liquid phase chromatography, supercritical fluid chromatography and Chromatography/Mass Spectrometry coupling technique; 3. immunological analysis method, mainly comprises enzyme linked immunosorbent assay analysis method, nm of gold immunity test strip method, fluoroimmunoassay etc.; 4. sensor method, mainly comprises piezocrystal biology sensor method, light transmitting fiber type enzyme sensor, organophosphor hydrolytic enzyme biology sensor, cholinesterase sensor and the biomimetic sensor based on molecular imprinting.5. fast detection method, mainly comprises quick measuring card method and tacheometer method.Spectroscopic methodology and chromatography be national authenticity detect the standard method of organophosphorus pesticide in agricultural product, there is high specificity, highly sensitive feature, its weak point is to need to utilize comparatively expensive instrument and special tester, and operating process is quite complicated, be not suitable for on-the-spot a large amount of organophosphorus pesticide in agricultural product that detects.Highly sensitive and the specificity of immunological detection method is stronger, but can only detect single or several organophosphorus pesticides, and the research and development time of corresponding antibodies is long and cost is high.Another shortcoming of immunological detection method is its repeatability and less stable.Sensor, method is the novel detection technique growing up in recent years, wherein about utilizing the current mode of cholinesterase and the report of electric potential type sensor aspect very many.The feature of sensing technology is sensitive, simple to operate, but achievement in research is still at laboratory stage, still exists the defect of repeatability and poor stability.Existing quick measuring card technology is to utilize cholinesterase can be hydrolyzed the analog 2 of acetylcholine, 6-dichloroindophenol acetic acid esters and generate blue 2,6-dichloroindophenol acetic acid, thus be determined with the residual condition of machine phosphorus insecticide according to the variation of quick measuring card color.Its advantage is that detection speed is fast, simple to operate.But the method is still simple quilitative method, and sensitivity is low, the organophosphorus to some kinds and the detectability of carbamate pesticide exceed the national limit standard of vegetables, fruit, and the organic pigment in sample is understood severe jamming mensuration and caused false negative result in addition.And existing tacheometer is still a kind of portable spectrophotometer, needs to coordinate comparatively complicated detection reagent, and be difficult at the scene implementing.
Find by prior art documents, number of patent application is 2005100741791.3, and name is called " chemiluminescent organophosphorus pesticide residual analyzer and detection method thereof ".Its gordian technique is, under the condition of potassium persulfate and the existence of nano titania ion, utilize ultraviolet catalytic effect to make the organophosphorus pesticide on vegetables or fruit be converted into inorganic phosphate, then form phosphorus molybdenum vanadium heteropolyacid with molybdic acid and vanadate, after this substance oxidation luminol, produce chemiluminescence, and measure with Chemiluminescence Apparatus.Be subject to extraneous phosphatic interference but its defect is reaction system, and make result occur false negative.The defect of this detection method is that repeatability, stability are still not ideal enough in addition, can not meet the needs of qualitative and quantitative detection.
Summary of the invention
The object of the invention is to the defect for prior art, a kind of kit and using method thereof of fast detecting organophosphorus pesticide is provided.
The present invention realizes by following technical scheme:
First aspect, the present invention relates to a kind of kit of fast detecting organophosphorus pesticide, described kit comprises: C enzyme bottle, T enzyme bottle, C substrate reactions cup, T substrate reactions cup, and described C enzyme bottle, T enzyme bottle are all used enzyme bottle seal cap sealing, and three enzyme intermixtures of enzyme freeze-drying are housed; Described C substrate reactions cup, T substrate reactions cup all arrange the freeze-drying adsorptive pads that contains acecoline, luminol and dimethylbenzidine.
Second aspect, the invention still further relates to the method for aforesaid kit fast detecting organophosphorus pesticide, comprises the following steps:
The 1st step: the preparation of enzyme cryoprotector: sucrose, bovine serum albumin(BSA), glycocoll, glycerine are added in calcium chloride solution, be stirred to completely and dissolve, freezing preservation is stand-by;
The 2nd step: the preparation of reaction enzymes liquid: described enzyme cryoprotector is diluted respectively to acetylcholinesterase, choline oxidase, peroxidase, mix, after freeze drying, be respectively charged in C enzyme bottle and T enzyme bottle;
Described cholinesterase is commercial acetylcholinesterase, and its source can be the acetylcholinesterase of producing from insect, animal's liver, fish brain, plant or technique for gene engineering, the acetylcholinesterase in the preferred insect of the application source.Described choline oxidase is also commercial choline oxidase, and it derives from the choline oxidase that Alcaligenes, soil Arthrobacter globiformis or technique for gene engineering are produced, the choline oxidase in the preferred Alcaligenes of the application source.Described peroxidase is commercially available horseradish peroxidase.
The 3rd step: the preparation of substrate reactions cup: reaction substrate pad is put into luminol, acecoline, dimethylbenzidine mixed solution, immersion drains, and after freeze drying, put into igelite reaction cup, be labeled as C substrate reactions cup, T substrate reactions cup;
The 4th step: the test of sample: drip pure water respectively in C, T enzyme bottle, after shaking up, in described T enzyme bottle, drip sample solution to be checked with plastic dropper, in C enzyme bottle, drip pure water or phosphate buffer, shake up, leave standstill, from described C, T enzyme bottle, draw reaction product with dropper, be added drop-wise to respectively in C, T substrate reactions cup, in 2～8 minutes, on portable chemical light-emitting appearance, measure the luminous intensity of C, T substrate reactions cup, according to the relative inhibition of luminous intensity, determine the concentration of organophosphorus.
Preferably, in the 1st step, enzyme cryoprotector comprises each component of following content: 100 milliliters of 5～15mmol/L calcium chloride solutions, 1～3 gram of sucrose, 1～3 gram of glycocoll, 5～15 milliliters of glycerine, 1～3 gram of bovine serum albumin(BSA).
Preferably, in the 2nd step, 3000～8000 times of described acetylcholinesterase dilutions, 200～800 times of described choline oxidase dilutions, 200～800 times of described peroxidase dilutions.
Preferably, in the 3rd step, the bottom surface internal diameter of described C substrate reactions cup, T substrate reactions cup is 8～20mm, is highly 10～25mm.
Preferably, in the 3rd step, the thickness of described reaction substrate pad is 3～6mm.
Preferably, in the 3rd step, the concentration of described luminol is 1.0 × 10
-4～3.0 × 10
-4the concentration of mol/L, acecoline is 3.0 × 10
-3～15.0 × 10
-3the concentration of mol/L, dimethylbenzidine is 2.0 × 10
-3～10.0 × 10
-3mol/L.
Preferably, in the 4th step, described is 50～200 microlitres to dripping pure water in C, T enzyme bottle, and described sample solution to be checked is 50～200 microlitres, and in described C enzyme bottle, pure water is 50～200 microlitres, and phosphate buffer is 0.1mol/L, and pH is 7.0.
Preferably, in the 4th step, reaction product 100～300 microlitres in described C, T substrate reactions cup.
The present invention has following beneficial effect:
(1) detection sensitivity is high: the present invention has selected three enzyme systems that contain acetylcholinesterase, choline oxidase and peroxidase, and set up the chemiluminescence based on sensitizer and luminol, its sensitivity exceeds decades of times and thousands of times, and detects the remarkable broadening of the range of linearity.The lowest detectable limit that the present invention detects DDVP and metrifonate reaches 0.078ppm; The lowest detectable limit that detects fenamiphos reaches 0.035ppm; Detectability to carbosulfan reaches 0.039ppm;
(2) kit of the present invention is easy to use, simple to operate, does not need complicated reagent preparation process in test, and layman just can complete detection easily according to kit instructions;
(3) detect fast: the present invention can complete pretreatment process and the testing process of sample within half an hour, and vegetables, fruit sample pre-treatments are only needed to 3～5 minutes, use kit test process to need 10～25 minutes.
(4) detectable organophosphorus scope is wide: the present invention can detect the several tens kinds of organophosphorus pesticides including Aldicarb, flolimat, Rogor, orthene, sevin, metrifonate, Furadan, carbosulfan, basudin, DDVP, Carbicron, cadusafos, ethyl parathion, fenamiphos, malathion, acephatemet, parathion-methyl, Azodrin, oxamyl, phosmet, phoxim, chlopyrifos, isocarbophos, the different sulphur phosphorus of methyl, azinphos-methyl etc.The present invention also can detect some heavy metals simultaneously and comprise residual on vegetables, fruit of the materials such as cadmium, copper, lead, mercury and zinc.
Brief description of the drawings
By reading the detailed description of non-limiting example being done with reference to the following drawings, it is more obvious that other features, objects and advantages of the present invention will become:
Fig. 1 is the mode chart of the kit of fast detecting organophosphorus pesticide of the present invention;
Wherein, 1 for containing three enzyme reagent bottle groups of contrast and test use, comprises C enzyme bottle, T enzyme bottle; 2 is enzyme bottle gland bonnet, and 3 is three enzyme reagent bottles; 4 is three enzyme intermixtures of enzyme freeze-drying; 5 is substrate reactions cup cup; 6 is the freeze-drying adsorptive pads that contains acecoline, luminol and dimethylbenzidine; 7 for contrast and test bigeminy substrate reactions cup group, comprises C substrate reactions cup, T substrate reactions cup.
Embodiment
Below in conjunction with specific embodiment, the present invention is described in detail.Following examples will contribute to those skilled in the art further to understand the present invention, but not limit in any form the present invention.It should be pointed out that to those skilled in the art, without departing from the inventive concept of the premise, can also make some distortion and improvement.These all belong to protection scope of the present invention.
embodiment 1
The present embodiment relates to a kind of method of utilizing kit fast detecting organophosphorus pesticide, comprises the following steps:
The present embodiment detects the residual of organophosphorus pesticide in commercially available rape.
1, sample pre-treatments, 10 parts of the commercially available rape leaves by weight within 10 grams, are positioned over one and are with in screw-topped plastics and glass container, also can be with conventional Cans.Add the pure water of approximately 10 milliliters, rock after 3 minutes, leave standstill after 2 minutes, get supernatant and can be directly used in test;
2, the preparation of enzyme cryoprotector: the compound method of the present embodiment is as follows: first prepare 100 milliliters of 5mmol/L calcium chloride solutions, then add 1 gram of sucrose, 1 gram of glycocoll, 5 milliliters of glycerine, 1 gram of bovine serum albumin(BSA), above composition is stirred while adding, until all powdered reagents all dissolve, then under-15 DEG C of conditions, freezing preservation is stand-by;
3, the preparation of reaction enzymes liquid: above-mentioned three kinds of enzyme liquid are diluted with above-mentioned enzyme cryoprotector; by 3000 times of every milligram of acetylcholinesterase dilutions that contains 425 international units of enzyme activity; by 200 times of every milligram of acetylcholinesterase dilutions that contains 10 international units of enzyme activity, by 200 times of every milligram of horseradish peroxidase dilutions that contains 113 international units of enzyme activity.Then respectively get above-mentioned enzyme liquid 100 microlitres and be mixed in together, be divided in brown ampoule bottle, be placed on freeze drier after freeze drying, after upper cover, seal, be positioned in aluminium foil bag room temperature preservation;
4, the preparation of substrate reactions cup: in the present embodiment, use igelite reaction cup, bottom surface internal diameter is 8mm, is highly 10mm, and the thickness of described reaction substrate pad is 3mm, puts into this pad that to contain concentration be 1.0 × 10
-4mol/L luminol, 3.0 × 10
-3mol/L acecoline and 2.0 × 10
-3in the mixed solution of mol/L dimethylbenzidine, soak and drain after 3 minutes, and after freeze drying, put into reaction cup, be labeled as respectively C and T cup, enclose in packaging bag, room temperature preservation is stand-by;
5, the test of sample: drip 50 microlitre pure water to C and T enzyme bottle respectively, after shaking up gently, to dripping 50 microlitre sample solution to be checked with quantitative plastic dropper in T enzyme bottle, to dripping 50 microlitre pure water with quantitative plastic dropper in C enzyme bottle, after shaking up gently, place 15 minutes, then draw reaction product 100 microlitres with dropper from the enzyme bottle of above-mentioned C and T, be added drop-wise to respectively in C and T substrate reactions cup, in 2 minutes, on portable chemical light-emitting appearance, measure the luminous intensity of C and T substrate reactions cup, according to the relative inhibition of luminous intensity, determine the concentration of organophosphorus.
Result shows, in 10 parts of tested rape samples, have the chemiluminescence inhibiting rate of 2 parts of rape samples to be respectively 16% and 33%, is judged to be the positive.The chemiluminescence inhibiting rate of all the other 8 parts of rape samples is all less than 15%.The flolimat that contrast test positive is 5ppm.
embodiment 2
The present embodiment relates to a kind of method of utilizing kit fast detecting organophosphorus pesticide, comprises the following steps:
The present embodiment detects the residual of organophosphorus pesticide in commercially available spinach.
1, sample pre-treatments, step is with the step 1 of embodiment 1;
2, the preparation of enzyme cryoprotector: the compound method of the present embodiment is as follows: first prepare 100 milliliters of 10mmol/L calcium chloride solutions, then add 2 grams of sucrose, 2 grams of glycocoll, 10 milliliters of glycerine, 2 grams of bovine serum albumin(BSA)s.Above composition is stirred while adding, until all powdered reagents all dissolve, then under-25 DEG C of conditions, freezing preservation is stand-by;
3, the preparation of reaction enzymes liquid: above-mentioned three kinds of enzyme liquid are diluted with above-mentioned enzyme cryoprotector; by 5000 times of every milligram of acetylcholinesterase dilutions that contains 425 international units of enzyme activity; by 500 times of every milligram of acetylcholinesterase dilutions that contains 10 international units of enzyme activity, by 500 times of every milligram of horseradish peroxidase dilutions that contains 113 international units of enzyme activity.Then respectively get above-mentioned enzyme liquid 200 microlitres and be mixed in together, be divided in brown ampoule bottle, be placed on freeze drier after freeze drying, after upper cover, seal, be positioned in aluminium foil bag room temperature preservation.
4, the preparation of substrate reactions cup: in the present embodiment, use polystyrene plastics reaction cup.Bottom surface internal diameter is 12mm, is highly 15mm.The thickness of described reaction substrate pad is 4mm.This pad is put into to contain concentration be 2.0 × 10
-4mol/L luminol, 8.0 × 10
-3mol/L acecoline and 6.0 × 10
-3in the mixed solution of mol/L dimethylbenzidine, soak and drain after 3 minutes, and after freeze drying, put into reaction cup, be labeled as respectively C and T cup, enclose in packaging bag, room temperature preservation is stand-by;
5, the test of sample: drip 100 microlitre pure water to C and T enzyme bottle respectively, after shaking up gently, to dripping 100 microlitre sample solution to be checked with quantitative plastic dropper in T enzyme bottle.To dripping 100 microlitre pure water with quantitative plastic dropper in C enzyme bottle, after shaking up gently, place 20 minutes.Then draw reaction product 200 microlitres with dropper from the enzyme bottle of above-mentioned C and T, be added drop-wise to respectively in C and T substrate reactions cup.Then in 5 minutes, on portable chemical light-emitting appearance, measure the luminous intensity of C and T substrate reactions cup, according to the relative inhibition of luminous intensity, determine the concentration of organophosphorus.
Result shows, in 10 parts of tested spinach samples, have the chemiluminescence inhibiting rate of 1 part of rape sample to be respectively 21%, is judged to be the positive.The chemiluminescence inhibiting rate of all the other 9 parts of spinach samples is all less than 15%.The Azodrin that contrast test positive is 10ppm.
embodiment 3
The present embodiment relates to a kind of method of utilizing kit fast detecting organophosphorus pesticide, comprises the following steps:
The present embodiment detects the residual of organophosphorus pesticide in commercially available crystal pears.
1, sample pre-treatments: first crystal pears are cut to 1/8th sizes, be positioned in clean Cans.All the other steps are with the step 1 of embodiment 1;
2, the preparation of enzyme cryoprotector: the compound method of the present embodiment is as follows: first prepare 100 milliliters of 15mmol/L calcium chloride solutions, then add 3 grams of sucrose, 3 grams of glycocoll, 15 milliliters of glycerine, 3 grams of bovine serum albumin(BSA)s.Above composition is stirred while adding, until all powdered reagents all dissolve, then under-40 DEG C of conditions, freezing preservation is stand-by;
3, the preparation of reaction enzymes liquid: above-mentioned three kinds of enzyme liquid are diluted with above-mentioned enzyme cryoprotector; by 8000 times of every milligram of acetylcholinesterase dilutions that contains 425 international units of enzyme activity; by 800 times of every milligram of acetylcholinesterase dilutions that contains 10 international units of enzyme activity, by 800 times of every milligram of horseradish peroxidase dilutions that contains 113 international units of enzyme activity.Then respectively get above-mentioned enzyme liquid 300 microlitres and be mixed in together, be divided in brown ampoule bottle, be placed on freeze drier after freeze drying, after upper cover, seal, be positioned in aluminium foil bag room temperature preservation.
4, the preparation of substrate reactions cup: in the present embodiment, use polycarbonate plastic reaction cup.Bottom surface internal diameter is 20mm, is highly 25mm.In the time of preparation feedback substrate pad.When described reaction substrate pad, thickness used is 6mm.Then this pad is put into to contain concentration be 3.0 × 10
-4mol/L luminol, 15.0 × 10
-3mol/L acecoline and 10.0 × 10
-3in the mixed solution of mol/L dimethylbenzidine, soak and drain after 3 minutes, and after freeze drying, put into reaction cup, be labeled as respectively C and T cup, enclose in packaging bag, room temperature preservation is stand-by;
5, the test of sample: drip 200 microlitre pure water to C and T enzyme bottle respectively, after shaking up gently, to dripping 200 microlitre sample solution to be checked with quantitative plastic dropper in T enzyme bottle.To dripping 200 microlitre pure water with quantitative plastic dropper in C enzyme bottle, after shaking up gently, place 25 minutes.Then draw reaction product 300 microlitres with dropper from the enzyme bottle of above-mentioned C and T, be added drop-wise to respectively in C and T substrate reactions cup.Then in 8 minutes, on portable chemical light-emitting appearance, measure the luminous intensity of C and T substrate reactions cup, according to the relative inhibition of luminous intensity, determine the concentration of organophosphorus.
Result shows, the chemiluminescence inhibiting rate of 10 parts of tested crystal pears samples is all less than 15%, and result is judged to be all negative.The acephatemet that contrast test positive is 15ppm.
The present invention utilizes acetylcholinesterase, choline oxidase and peroxidase for catalyst system and catalyzing, and the product after acecoline hydrolysis oxidation and luminol are produced to chemiluminescence effect.Because organophosphorus pesticide can suppress the activity of three kinds of enzymes of this reaction system, especially the activity of acetylcholinesterase, thereby can set up the method for testing delicately organophosphorus pesticide in agricultural product, and utilize this principle to develop the chemiluminescence detection kit that detects organophosphorus pesticide.The present invention has that tool detection speed is fast, operating process is simple, stability and reproducible, do not need the advantages such as expensive instrument, be very suitable for carrying out in the market of farm produce field monitoring of organophosphorus pesticide.
Above specific embodiments of the invention are described.It will be appreciated that, the present invention is not limited to above-mentioned specific implementations, and those skilled in the art can make various distortion or amendment within the scope of the claims, and this does not affect flesh and blood of the present invention.
Claims (9)
1. the kit of a fast detecting organophosphorus pesticide, it is characterized in that, described kit comprises: C enzyme bottle, T enzyme bottle, C substrate reactions cup, T substrate reactions cup, and described C enzyme bottle, T enzyme bottle are all used enzyme bottle seal cap sealing, and three enzyme intermixtures of enzyme freeze-drying are housed; Described C substrate reactions cup, T substrate reactions cup all arrange the freeze-drying adsorptive pads that contains acecoline, luminol and dimethylbenzidine.
2. application rights requires a method for the kit fast detecting organophosphorus pesticide described in 1, it is characterized in that, comprises the following steps:
The 1st step: the preparation of enzyme cryoprotector: sucrose, bovine serum albumin(BSA), glycocoll, glycerine are added in calcium chloride solution, be stirred to completely and dissolve, freezing preservation is stand-by;
The 2nd step: the preparation of reaction enzymes liquid: described enzyme cryoprotector is diluted respectively to acetylcholinesterase, choline oxidase, peroxidase, mix, after freeze drying, be respectively charged in C enzyme bottle and T enzyme bottle;
The 3rd step: the preparation of substrate reactions cup: reaction substrate pad is put into luminol, acecoline, dimethylbenzidine mixed solution, immersion drains, and after freeze drying, put into igelite reaction cup, be labeled as C substrate reactions cup, T substrate reactions cup;
The 4th step: the test of sample: drip pure water respectively in C, T enzyme bottle, after shaking up, in described T enzyme bottle, drip sample solution to be checked with plastic dropper, in C enzyme bottle, drip pure water or phosphate buffer, shake up, leave standstill, from described C, T enzyme bottle, draw reaction product with dropper, be added drop-wise to respectively in C, T substrate reactions cup, in 2～8 minutes, on portable chemical light-emitting appearance, measure the luminous intensity of C, T substrate reactions cup, according to the relative inhibition of luminous intensity, determine the concentration of organophosphorus.
3. the method for kit fast detecting organophosphorus pesticide as claimed in claim 2; it is characterized in that; in the 1st step; enzyme cryoprotector comprises each component of following content: 100 milliliters of 5～15mmol/L calcium chloride solutions; 1～3 gram of sucrose; 1～3 gram of glycocoll, 5～15 milliliters of glycerine, 1～3 gram of bovine serum albumin(BSA).
4. the method for kit fast detecting organophosphorus pesticide as claimed in claim 2, it is characterized in that, in the 2nd step, 3000～8000 times of described acetylcholinesterase dilutions, 200～800 times of described choline oxidase dilutions, 200～800 times of described peroxidase dilutions.
5. the method for kit fast detecting organophosphorus pesticide as claimed in claim 2, is characterized in that, in the 3rd step, the bottom surface internal diameter of described C substrate reactions cup, T substrate reactions cup is 8～20mm, is highly 10～25mm.
6. the method for kit fast detecting organophosphorus pesticide as claimed in claim 2, is characterized in that, in the 3rd step, the thickness of described reaction substrate pad is 3～6mm.
7. the method for kit fast detecting organophosphorus pesticide as claimed in claim 2, is characterized in that, in the 3rd step, the concentration of described luminol is 1.0 × 10
-4～3.0 × 10
-4the concentration of mol/L, acecoline is 3.0 × 10
-3～15.0 × 10
-3the concentration of mol/L, dimethylbenzidine is 2.0 × 10
-3～10.0 × 10
-3mol/L.
8. the method for kit fast detecting organophosphorus pesticide as claimed in claim 2, it is characterized in that, in the 4th step, described is 50～200 microlitres to dripping pure water in C, T enzyme bottle, described sample solution to be checked is 50～200 microlitres, in described C enzyme bottle, pure water is 50～200 microlitres, and phosphate buffer is 0.1mol/L, and pH is 7.0.
9. the method for kit fast detecting organophosphorus pesticide as claimed in claim 2, is characterized in that, in the 4th step, and reaction product 100～300 microlitres in described C, T substrate reactions cup.
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| CN105241882A (en) * | 2015-09-23 | 2016-01-13 | 山东大学 | Applications of liquid crystal sensor in detection of organophosphorus pesticides and carbamate pesticides |
| CN105319211A (en) * | 2015-11-13 | 2016-02-10 | 无锡艾科瑞思产品设计与研究有限公司 | Kit for quickly detecting organophosphorus residues in agricultural products and application method of kit |
| CN105548431B (en) * | 2015-12-14 | 2016-09-07 | 山东省农业科学院植物保护研究所 | Detect the method for oxamyl and oxamyl oxime residual quantity in vegetables/fruit simultaneously |
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| CN114295769A (en) * | 2021-12-28 | 2022-04-08 | 云南省农业科学院质量标准与检测技术研究所 | Method for rapidly identifying recessive addition of forbidden pesticides in fertilizer |
| CN115595318A (en) * | 2022-09-22 | 2023-01-13 | 大连理工大学（Cn） | Method for ZIF-8 in-situ immobilized enzyme with high enzyme activity and application thereof |
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