CN106568946A - immunogold detection card of propiconazole, and preparation method thereof - Google Patents
immunogold detection card of propiconazole, and preparation method thereof Download PDFInfo
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- CN106568946A CN106568946A CN201510656041.0A CN201510656041A CN106568946A CN 106568946 A CN106568946 A CN 106568946A CN 201510656041 A CN201510656041 A CN 201510656041A CN 106568946 A CN106568946 A CN 106568946A
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- propiconazole
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/558—Immunoassay; Biospecific binding assay; Materials therefor using diffusion or migration of antigen or antibody
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/531—Production of immunochemical test materials
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/531—Production of immunochemical test materials
- G01N33/532—Production of labelled immunochemicals
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2430/00—Assays, e.g. immunoassays or enzyme assays, involving synthetic organic compounds as analytes
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- Urology & Nephrology (AREA)
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- Food Science & Technology (AREA)
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- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
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- Investigating Or Analysing Biological Materials (AREA)
- Peptides Or Proteins (AREA)
Abstract
The invention provides an immunogold detection card of propiconazole, and a preparation method thereof, and relates to the technical field of animal derived food veterinary drug residue detection. According to the detection card, the test paper strip in the shell comprises a PVC glue plate, a sample pad, a colloidal gold binding pad, a coating film and a water absorption pad, wherein the colloidal gold film is a glass cellulose film containing propiconazole monoclonal antibody, the coating film is a nitrocellulose film and is provided with a T line and a C line, the T line is coated with a propiconazole protein conjugate, and the C line is coated with goat anti-mouse IgG antibody. The immunogold detection card of the present invention can be effectively used for rapid and convenient detection of propiconazole, wherein the result is accurate.
Description
Technical field
The present invention relates to immune colloid gold detection card of cereal foods Pesticide Residues detection technique field, more particularly to propiconazole and preparation method thereof.
Background technology
Propiconazole（propiconazole）It is a kind of absorbability triazole type New-type wide-spectrum antibacterial with treatment and protection dual function; can be absorbed by root, stem, leaf portion; and soon can conduct upwards in plant strain body; the disease that preventing and treating ascomycetess, basidiomycetes and Fungi Imperfecti cause; particularly there is specially good effect to diseases such as take-all, powdery mildew, rust, root rot, bakanae disease of rice, banded sclerotial blight, sigatokas; the disease that most of higher funguses cause can be effectively prevented and treated, but it is invalid to oomycetes disease.
Prior art is mainly the detection methods such as gas chromatography-mass spectrography, high performance liquid chromatography for the detection of propiconazole, but the defect of these technologies is obvious：Apparatus expensive, complex operation, it is difficult to promote.So, set up it is a kind of it is simple, effectively, to be adapted to the assay method that basic unit uses be extremely necessary.The purpose of the present invention is to develop a kind of enterprise that is more suitable for carry out Site Detection and qualitative checking method fast and simple, with low cost.
The content of the invention
For case above, the purpose of the present invention is exactly the defect in order to overcome prior art to exist and provides immune colloid gold detection card of a kind of propiconazole and preparation method thereof, can effectively solving can quick, easily detect the problem of propiconazole.
The propiconazole colloidal-gold detecting-card of the present invention, constitute including the gold conjugation pad for being coated with monoclonal antibody colloid gold label thing, the nitrocellulose filter, sample pad, adsorptive pads, PVC offset plates and the mould of plastics that have been coated with propiconazole-BSA and sheep anti-mouse igg, sample pad, pad are adhered to successively in one end of PVC offset plates, paste nitrocellulose filter, other end adhesion adsorptive pads in centre.
The preparation method of the propiconazole colloidal-gold detecting-card of the present invention, is realized by following steps：
(1) prepared by colloidal gold solution：Take 1
L conical flasks 1, add the mL of ultra-pure water 495, then add 1% gold chloride (HAuCl4·3H2O)
5 mL, are configured to the aqueous solution of chloraurate of 500 mL 0.01%, and 1% trisodium citrate (Na is added in the case where persistently stirring after heated and boiled3C6H5O7·2H2O) solution 5-7 mL, continue agitating heating, when the color of solution is changed into completely transparent aubergine, maintain to stop heating after 5 min, and moisturizing is cooled to room temperature to original volume, and 2-8 DEG C saves backup；
(2) pretreatment of antibody：Will labelling propiconazole antibody in 1000 r/min, under the conditions of 4 DEG C, be centrifuged 20 min, take supernatant, be diluted to 1 with 0.01 mol/L PBS
mg/mL；Or 1 mg/ml is diluted to 0.01 mol/L PBS, cross 0.22 m filter membranes；
(3) preparation of colloid gold label thing：The mL of colloidal gold solution 40 in step (1) is taken, with 0.25 mol/L K2CO3 colloidal gold solution pH to 8.5, magnetic stirrer 250 are adjusted
R/min is stirred, and is added dropwise over protein solutions of 4 mL containing 0.3 mg antibody proteins, reacts 10 min；It is added dropwise over 4 mL
10% BSA, continues the min of stirring reaction 10；Gold labeling antibody solution room temperature low speed（1500 r/min）10 min are centrifuged, the precipitation formed by the gold grain for condensing is discarded；4 DEG C of red supernatant solution, 12000 r/min are centrifuged 20 min, abandon supernatant, collect precipitation, and precipitation gold labeling antibody diluent is settled to into 1 mL, are prepared into propiconazole monoclonal antibody colloid gold label thing；
(4) preparation of colloidal gold film：Step (3) propiconazole protein conjugate monoclonal antibody colloid gold label thing is sprayed on carrier glass cellulose membrane with film instrument is drawn with the even concentration of 5 L/cm, room temperature is dried or 37 DEG C of 3 h of drying naturally, makes the colloidal gold film of the monoclonal antibody colloid gold label thing of protein conjugate containing propiconazole；
(5) it is coated with film preparation：Sheep anti-mouse igg antibody, propiconazole protein conjugate are diluted to into 1 mg/mL, are sprayed on nitrocellulose filter with the concentration of 1 L/cm successively with film instrument is drawn, be prepared into coated film, after 37 DEG C of 2 h of coating, room temperature is dried or 37 DEG C of drying naturally；
(6) sample pad pre-treatment：Sample pad treatment fluid is uniformly coated on glass fibre element film, is less than under conditions of 60% in air humidity, room temperature is dried naturally；
(7) assembling of detection card：PVC offset plates paste successively from top to bottom sample pad, colloidal gold film, coated film and absorbent cotton, are assembled into test strips, cut into the strip of one fixed width, then test strips are arranged in the detection card shell of the flat shelly of strip.
In the step (5) coated detection line T be located at the lower section of nature controlling line C；
Sample pad treatment fluid in the step (6) is by 1 g bovin serum albumin (BSA) and 0.8 g Sodium Chloride (NaCl), with containing 0.5%TRITON-100 0.01
Mol/L PBS are settled to 100 mL；
The present invention can be effectively used for determining propiconazole, and method is simple, convenient, fast, as a result accurately.
Description of the drawings
Fig. 1 is the structure chart of the immune colloid gold detection card of propiconazole of the present invention：1. well in figure
2. the detection of 3. nature controlling line of detection line 4. 5. test strips of hole 6. detect card shell.
Fig. 2 be propiconazole of the present invention immune colloid gold detection card in test strips sectional structure chart, 7. sample pad in figure
8. adsorptive pads of 10. nitrocellulose filter of gold conjugation pad 9.PVC offset plates 11..
Specific embodiment
Embodiment 1
Embodiment shown in Fig. 1, Fig. 2：9 is PVC offset plates in figure；7 is sample pad；8 is gold conjugation pad, and monoclonal antibody colloid gold label thing has been coated with the gold conjugation pad；10 is coated film, i.e. nitrocellulose filter, and propiconazole-BSA and sheep anti-mouse igg have been coated with the nitrocellulose filter；11 is adsorptive pads, is made up of absorbent material such as filter paper.
(sample end) adhesion sample pad 7, pad 8 on one end of PVC offset plates 9, sample pad 7 and pad 8 are side by side configuration.
In the intermediate adhesion nitrocellulose filter 10 of PVC offset plates 9.Sheep anti-mouse igg nature controlling line 3 and propiconazole-BSA detection lines 2 are provided with nitrocellulose filter 10.
Adsorptive pads 11 are adhered in the other end of PVC offset plates 9.One end of nitrocellulose filter 10 slightly intersects with pad 8, and the other end slightly intersects with adsorptive pads 11.The test strips 5 can be incorporated with made by mould of plastics and detect in card shell 6, make detection card, and well 1 and detection hole 4 are provided with the upper lid of detection card shell 6, and, just to well 1, nitrocellulose filter 10 is just to detecting hole 4 for sample pad 7.
Embodiment 2
Prepared by propiconazole colloidal-gold detecting-card, implemented by following steps：
It is prepared by colloidal gold solution：1 L conical flasks 1 are taken, the mL of ultra-pure water 495 is added, 1% gold chloride (HAuCl is then added4·3H2O)
5 mL, are configured to the aqueous solution of chloraurate of 500 mL 0.01%, and 1% trisodium citrate (Na is added in the case where persistently stirring after heated and boiled3C6H5O7·2H2O) solution 5-7 mL, continue agitating heating, when the color of solution is changed into completely transparent aubergine, maintain to stop heating after 5 min, and moisturizing is cooled to room temperature to original volume, and 2-8 DEG C saves backup；
The pretreatment of antibody：Will labelling propiconazole antibody in 1000 r/min, under the conditions of 4 DEG C, centrifugation 20
Min, takes supernatant, and with 0.01 mol/L PBS 1 mg/mL is diluted to；Or use 0.01
Mol/L PBS are diluted to 1 mg/ml, cross 0.22 m filter membranes；
The preparation of colloid gold label thing：The mL of colloidal gold solution 40 in step (1) is taken, with 0.25 mol/L K2CO3 colloidal gold solution pH to 8.5, magnetic stirrer 250 are adjusted
R/min is stirred, and is added dropwise over protein solutions of 4 mL containing 0.3 mg antibody proteins, reacts 10 min；It is added dropwise over 4 mL
10% BSA, continues the min of stirring reaction 10；Gold labeling antibody solution room temperature low speed（1500 r/min）10 min are centrifuged, the precipitation formed by the gold grain for condensing is discarded；4 DEG C of red supernatant solution, 12000 r/min are centrifuged 20 min, abandon supernatant, collect precipitation, and precipitation gold labeling antibody diluent is settled to into 1 mL, are prepared into propiconazole monoclonal antibody colloid gold label thing；
The preparation of colloidal gold film：Step (3) propiconazole protein conjugate monoclonal antibody colloid gold label thing is sprayed on carrier glass cellulose membrane with film instrument is drawn with the even concentration of 5 L/cm, room temperature is dried or 37 DEG C of 3 h of drying naturally, makes the colloidal gold film of the monoclonal antibody colloid gold label thing of protein conjugate containing propiconazole；
Coating film preparation：Sheep anti-mouse igg antibody, propiconazole protein conjugate are diluted to into 1 mg/mL, with stroke film instrument successively with 1
The concentration of L/cm is sprayed on nitrocellulose filter, is prepared into coated film, and after 37 DEG C of 2 h of coating, room temperature is dried or 37 DEG C of drying naturally；
Sample pad pre-treatment：Sample pad treatment fluid is uniformly coated on glass fibre element film, is less than under conditions of 60% in air humidity, room temperature is dried naturally；
The assembling of detection card：PVC offset plates paste successively from top to bottom sample pad, colloidal gold film, coated film and absorbent cotton, are assembled into test strips, cut into one fixed width strip, then test strips are arranged in the detection card shell of the flat shelly of strip.
Claims (2)
1. propiconazole colloidal-gold detecting-card, it is characterised in that prepare as steps described below：
(1) prepared by colloidal gold solution：1 L conical flasks 1 are taken, the mL of ultra-pure water 495 is added, 1% gold chloride HAuCl is then added4·3H2O) 5 mL, is configured to the aqueous solution of chloraurate of 500 mL 0.01%, and 1% trisodium citrate Na is added in the case where persistently stirring after heated and boiled3C6H5O7·2H2O solution 5-7 mL, continue agitating heating, when the color of solution is changed into completely transparent aubergine, maintain to stop heating after 5 min, and moisturizing is cooled to room temperature to original volume, and 2-8 DEG C saves backup；
(2) pretreatment of antibody：Will labelling propiconazole antibody 1000
R/min, under the conditions of 4 DEG C, is centrifuged 20 min, takes supernatant, and with 0.01 mol/L PBS 1 mg/mL is diluted to；Or 1 mg/ml is diluted to 0.01 mol/L PBS, cross 0.22 m filter membranes；
(3) preparation of colloid gold label thing：The mL of colloidal gold solution 40 in step (1) is taken, with 0.25 mol/L K2CO3Adjust colloidal gold solution pH to 8.5, magnetic stirrer 250
R/min is stirred, and is added dropwise over protein solutions of 4 mL containing 0.3 mg antibody proteins, reacts 10 min；The BSA of 4 mL 10% are added dropwise over, continue the min of stirring reaction 10；The r/min of gold labeling antibody solution room temperature 1500 is centrifuged 10 min, discards the precipitation formed by the gold grain for condensing；4 DEG C of red supernatant solution, 12000 r/min are centrifuged 20 min, abandon supernatant, collect precipitation, and precipitation gold labeling antibody diluent is settled to into 1 mL, are prepared into propiconazole monoclonal antibody colloid gold label thing；
(4) preparation of colloidal gold film：Step (3) propiconazole protein conjugate monoclonal antibody colloid gold label thing is sprayed on carrier glass cellulose membrane with film instrument is drawn with the even concentration of 5 L/cm, room temperature is dried or 37 DEG C of 3 h of drying naturally, makes the colloidal gold film of the monoclonal antibody colloid gold label thing of protein conjugate containing propiconazole；
(5) it is coated with film preparation：Sheep anti-mouse igg antibody, propiconazole protein conjugate are diluted to into 1 mg/mL, are sprayed on nitrocellulose filter with the concentration of 1 L/cm successively with film instrument is drawn, be prepared into coated film, after 37 DEG C of 2 h of coating, room temperature is dried or 37 DEG C of drying naturally；
(6) sample pad pre-treatment：Sample pad treatment fluid is uniformly coated on glass fibre element film, is less than under conditions of 60% in air humidity, room temperature is dried naturally；
(7) assembling of detection card：PVC offset plates paste successively from top to bottom sample pad, colloidal gold film, coated film and absorbent cotton, are assembled into test strips, cut into the strip of one fixed width, then test strips are arranged in the detection card shell of the flat shelly of strip.
2. propiconazole colloidal-gold detecting-card according to claim 1, it is characterised in that in the step (5) coated detection line T be located at the lower section of nature controlling line C；
Sample pad treatment fluid in the step (6) is, by 1 g bovin serum albumin (BSA) and 0.8 g Sodium Chloride (NaCl), with the 0.01 mol/L PBS containing 0.5%TRITON-100 100 mL to be settled to.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510656041.0A CN106568946A (en) | 2015-10-13 | 2015-10-13 | immunogold detection card of propiconazole, and preparation method thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510656041.0A CN106568946A (en) | 2015-10-13 | 2015-10-13 | immunogold detection card of propiconazole, and preparation method thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN106568946A true CN106568946A (en) | 2017-04-19 |
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| Application Number | Title | Priority Date | Filing Date |
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| CN201510656041.0A Withdrawn CN106568946A (en) | 2015-10-13 | 2015-10-13 | immunogold detection card of propiconazole, and preparation method thereof |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109813891A (en) * | 2017-11-18 | 2019-05-28 | 镇江亿特生物科技发展有限公司 | A kind of time-resolved fluoroimmunoassay chromatography quantitative testing test paper item detecting chlorpromazine |
| CN109813901A (en) * | 2017-11-21 | 2019-05-28 | 南京亿特生物科技有限公司 | A kind of immune colloid gold detection card and preparation method thereof detecting ethiprole |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102901819A (en) * | 2012-09-27 | 2013-01-30 | 江苏维赛科技生物发展有限公司 | Immune colloidal gold detection card for phenylethanolamine A and preparation method thereof |
| CN103808930A (en) * | 2012-11-06 | 2014-05-21 | 江苏维赛科技生物发展有限公司 | Zilpaterol immune colloidal gold detection card and preparation method thereof |
| CN104655835A (en) * | 2013-11-20 | 2015-05-27 | 南京亿特生物科技有限公司 | Immune colloidal gold detect card of chlorpromazine, and preparation method thereof |
-
2015
- 2015-10-13 CN CN201510656041.0A patent/CN106568946A/en not_active Withdrawn
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102901819A (en) * | 2012-09-27 | 2013-01-30 | 江苏维赛科技生物发展有限公司 | Immune colloidal gold detection card for phenylethanolamine A and preparation method thereof |
| CN103808930A (en) * | 2012-11-06 | 2014-05-21 | 江苏维赛科技生物发展有限公司 | Zilpaterol immune colloidal gold detection card and preparation method thereof |
| CN104655835A (en) * | 2013-11-20 | 2015-05-27 | 南京亿特生物科技有限公司 | Immune colloidal gold detect card of chlorpromazine, and preparation method thereof |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109813891A (en) * | 2017-11-18 | 2019-05-28 | 镇江亿特生物科技发展有限公司 | A kind of time-resolved fluoroimmunoassay chromatography quantitative testing test paper item detecting chlorpromazine |
| CN109813901A (en) * | 2017-11-21 | 2019-05-28 | 南京亿特生物科技有限公司 | A kind of immune colloid gold detection card and preparation method thereof detecting ethiprole |
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Application publication date: 20170419 |
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