CN105572373A - Immune colloidal gold detecting card of carbendazin and preparation method of immune colloidal gold detecting card - Google Patents
Immune colloidal gold detecting card of carbendazin and preparation method of immune colloidal gold detecting card Download PDFInfo
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- CN105572373A CN105572373A CN201410551852.XA CN201410551852A CN105572373A CN 105572373 A CN105572373 A CN 105572373A CN 201410551852 A CN201410551852 A CN 201410551852A CN 105572373 A CN105572373 A CN 105572373A
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Abstract
The invention discloses an immune colloidal gold detecting card of carbendazin and a preparation method of the immune colloidal gold detecting card, and relates to the technical field of animal source food veterinary drug residue detection. A test strip in a shell of the detecting card is composed of a PVC glue plate, a sample cushion, a colloidal gold conjugate pad, a coating film and a water absorbing cushion, wherein a colloidal gold film is a glass cellulose membrane containing carbendazin monoclonal antibodies, the coating film is a nitrocellulose film and is provided with a line T and a line C, the line T is wrapped by carbendazin protein conjugates, and the line C is wrapped by goat-anti-mouse IgG antibodies. The detecting card can be effectively used for rapidly detecting carbendazin and is convenient to use, rapid and accurate in result.
Description
Technical field
The present invention relates to cereal foods Pesticide Residues detection technique field, immune colloid gold test card particularly relating to carbendazim and preparation method thereof.
Background technology
Carbendazim (Carbendazin), chemical name 2-(methoxyformamido) benzimidazole [2-(methoxy-carbanmoyl)-benzimidaxole], be a kind of high-efficiency low-toxicity germifuge, can be used for preventing and treating plurality of plant diseases.Carbendazim is a kind of agricultural chemicals conventional in asparagus cultivation process, and China's asparagus products export amount is very large, and residues of pesticides are very important restriction indexs.Therefore regularly carbendazim is remained to the necessary means detecting the monitoring becoming many countries.European Union specifies that the maximum residue limit(MRL) of carbendazim in vegetables (MRL) is 0.1mg/kg.
Prior art is for the detection mainly detection method such as gas chromatography-mass spectrography, high performance liquid chromatography of carbendazim, but the defect of these technology is obvious: apparatus expensive, complicated operation, be difficult to promote.So it is extremely necessary for setting up a kind of assay method that is simple, effective, that be applicable to basic unit's use.The object of the invention is a kind of enterprise that is more suitable for of development and carry out Site Detection and qualitative checking method fast and simple, with low cost.
Summary of the invention
For above situation, object of the present invention be exactly in order to overcome prior art exist defect and immune colloid gold test card that a kind of carbendazim is provided and preparation method thereof, effectively can solve the problem that can detect carbendazim fast, easily.
Carbendazim colloidal-gold detecting-card of the present invention, comprise and being formed by the nitrocellulose filter of carbendazim-BSA and sheep anti-mouse igg, sample pad, adsorptive pads, PVC offset plate and mould of plastics by the gold conjugation pad of monoclonal antibody colloid gold label thing, bag, sample pad, pad is adhered to successively in one end of PVC offset plate, nitrocellulose filter is pasted in centre, and the other end adheres to adsorptive pads.
The preparation method of carbendazim colloidal-gold detecting-card of the present invention is realized by following steps:
(1) colloidal gold solution preparation: get 1L Erlenmeyer flask 1, add ultrapure water 495mL, then add 1% gold chloride (HAuCl
43H
2o) 5mL, is mixed with 500mL0.01% aqueous solution of chloraurate, and heating adds 1% trisodium citrate (Na after boiling when Keep agitation
3c
6h
5o
72H
2o) solution 5-7mL, continues agitating heating, and when the color of solution becomes transparent aubergine completely, stop heating after maintaining 5min, moisturizing, to original volume, is cooled to room temperature, and 2-8 DEG C saves backup;
(2) pre-service of antibody: the carbendazim antibody that will mark is at 1000r/min, and under 4 DEG C of conditions, centrifugal 20min, gets supernatant, is diluted to 1mg/mL with 0.01mol/LPBS; Or be diluted to 1mg/ml with 0.01mol/LPBS, cross 0.22 μm of filter membrane;
(3) preparation of colloid gold label thing: get the colloidal gold solution 40mL in step (1), colloidal gold solution pH to 8.5 is regulated with 0.25mol/LK2CO3, magnetic stirrer 250r/min stirs, and dropwise adds the protein solution of 4mL containing 0.3mg antibody protein, reaction 10min; Dropwise add 4mL10%BSA, continue stirring reaction 10min; Gold labeling antibody solution normal temperature low speed (1500r/min) centrifugal 10min, discards the precipitation formed by the gold grain condensed; Red supernatant solution 4 DEG C, the centrifugal 20min of 12000r/min, abandons supernatant, collecting precipitation, and the golden labeling antibody dilution of precipitation is settled to 1mL, is prepared into carbendazim monoclonal antibody colloid gold label thing;
(4) preparation of colloidal gold film: step (3) carbendazim protein conjugate monoclonal antibody colloid gold label thing is sprayed on carrier glass cellulose membrane with drawing film instrument with the even concentration of 5 μ L/cm, room temperature is dried or 37 DEG C of oven dry 3h naturally, makes the colloidal gold film containing carbendazim protein conjugate monoclonal antibody colloid gold label thing;
(5) coated film preparation: sheep anti-mouse igg antibody, carbendazim protein conjugate are diluted to 1mg/mL, be sprayed on nitrocellulose filter with the concentration of 1 μ L/cm successively with drawing film instrument, be prepared into coated film, 37 DEG C of bags are by after 2h, and room temperature is dried or 37 DEG C of oven dry naturally;
(6) sample pad pre-treatment: be uniformly coated on by sample pad treating fluid on glass fibre element film, under the condition of air humidity lower than 60%, room temperature is dried naturally;
(7) assembling of test card: PVC offset plate pastes sample pad, colloidal gold film, coated film and absorbent wool from top to bottom successively, is assembled into test strips, cuts into the rectangular of one fixed width, then test strips is arranged in the test card shell of rectangular flat shelly.
In described step (5), the detection line T of institute's bag quilt is located at the below of nature controlling line C;
Sample pad treating fluid in described step (6) is by 1g Bovine serum albumin (BSA) and 0.8g sodium chloride (NaCl), is settled to 100mL with the 0.01mol/LPBS containing 0.5%TRITON-100;
The present invention can be effective to measure carbendazim, and method is simple, and convenient, fast, result is accurate.
Accompanying drawing explanation
Fig. 1 is the structural drawing of the immune colloid gold test card of carbendazim of the present invention: 1. well 2. detection line 3. nature controlling line 4. detect aperture 5. test strips 6. test card shells in figure.
Fig. 2 is the sectional structure chart of the test strips in the immune colloid gold test card of carbendazim of the present invention, 7. sample pad 8. gold conjugation pad 9.PVC offset plate 10. nitrocellulose filter 11. adsorptive pads in figure.
Embodiment
Embodiment 1
Embodiment shown in Fig. 1, Fig. 2: in figure, 9 is PVC offset plate; 7 is sample pad; 8 is gold conjugation pad, and on this gold conjugation pad, bag is by monoclonal antibody colloid gold label thing; 10 is coated film, i.e. nitrocellulose filter, and on this nitrocellulose filter, bag is by carbendazim-BSA and sheep anti-mouse igg; 11 is adsorptive pads, by absorbent material as filter paper is made.
On one end of PVC offset plate 9, (sample end) adheres to sample pad 7, pad 8, and sample pad 7 and pad 8 are side by side configuration.
At the intermediate adhesion nitrocellulose filter 10 of PVC offset plate 9.Nitrocellulose filter 10 is provided with sheep anti-mouse igg nature controlling line 3 and carbendazim-BSA detection line 2.
Adsorptive pads 11 is adhered at the other end of PVC offset plate 9.One end of nitrocellulose filter 10 slightly intersects with pad 8, and the other end slightly intersects with adsorptive pads 11.This test strips 5 can be incorporated with in the test card shell 6 that mould of plastics makes, and make test card, test card shell 6 covers and is provided with well 1 and detect aperture 4, sample pad 7 is just to well 1, and nitrocellulose filter 10 is just to detect aperture 4.
Embodiment 2
Prepared by carbendazim colloidal-gold detecting-card, be by following steps specific implementation:
Prepared by colloidal gold solution: get 1L Erlenmeyer flask 1, add ultrapure water 495mL, then add 1% gold chloride (HAuCl
43H
2o) 5mL, is mixed with 500mL0.01% aqueous solution of chloraurate, and heating adds 1% trisodium citrate (Na after boiling when Keep agitation
3c
6h
5o
72H
2o) solution 5-7mL, continues agitating heating, and when the color of solution becomes transparent aubergine completely, stop heating after maintaining 5min, moisturizing, to original volume, is cooled to room temperature, and 2-8 DEG C saves backup;
The pre-service of antibody: the carbendazim antibody that will mark is at 1000r/min, and under 4 DEG C of conditions, centrifugal 20min, gets supernatant, is diluted to 1mg/mL with 0.01mol/LPBS; Or be diluted to 1mg/ml with 0.01mol/LPBS, cross 0.22 μm of filter membrane;
The preparation of colloid gold label thing: get the colloidal gold solution 40mL in step (1), colloidal gold solution pH to 8.5 is regulated with 0.25mol/LK2CO3, magnetic stirrer 250r/min stirs, and dropwise adds the protein solution of 4mL containing 0.3mg antibody protein, reaction 10min; Dropwise add 4mL10%BSA, continue stirring reaction 10min; Gold labeling antibody solution normal temperature low speed (1500r/min) centrifugal 10min, discards the precipitation formed by the gold grain condensed; Red supernatant solution 4 DEG C, the centrifugal 20min of 12000r/min, abandons supernatant, collecting precipitation, and the golden labeling antibody dilution of precipitation is settled to 1mL, is prepared into carbendazim monoclonal antibody colloid gold label thing;
The preparation of colloidal gold film: step (3) carbendazim protein conjugate monoclonal antibody colloid gold label thing is sprayed on carrier glass cellulose membrane with drawing film instrument with the even concentration of 5 μ L/cm, room temperature is dried or 37 DEG C of oven dry 3h naturally, makes the colloidal gold film containing carbendazim protein conjugate monoclonal antibody colloid gold label thing;
Prepared by coated film: sheep anti-mouse igg antibody, carbendazim protein conjugate are diluted to 1mg/mL, be sprayed on nitrocellulose filter with the concentration of 1 μ L/cm successively with drawing film instrument, be prepared into coated film, 37 DEG C of bags are by after 2h, and room temperature is dried or 37 DEG C of oven dry naturally;
Sample pad pre-treatment: be uniformly coated on by sample pad treating fluid on glass fibre element film, under the condition of air humidity lower than 60%, room temperature is dried naturally;
The assembling of test card: PVC offset plate pastes sample pad, colloidal gold film, coated film and absorbent wool from top to bottom successively, is assembled into test strips, cuts into one fixed width rectangular, then test strips is arranged in the test card shell of rectangular flat shelly.
Claims (2)
1. carbendazim colloidal-gold detecting-card, is characterized in that preparing according to following step:
(1) colloidal gold solution preparation: get 1L Erlenmeyer flask 1, add ultrapure water 495mL, then add 1% gold chloride HAuCl
43H
2o) 5mL, is mixed with 500mL0.01% aqueous solution of chloraurate, and heating adds 1% trisodium citrate Na after boiling when Keep agitation
3c
6h
5o
72H
2o solution 5-7mL, continues agitating heating, and when the color of solution becomes transparent aubergine completely, stop heating after maintaining 5min, moisturizing, to original volume, is cooled to room temperature, and 2-8 DEG C saves backup;
(2) pre-service of antibody: the carbendazim antibody that will mark is at 1000r/min, and under 4 DEG C of conditions, centrifugal 20min, gets supernatant, is diluted to 1mg/mL with 0.01mol/LPBS; Or be diluted to 1mg/ml with 0.01mol/LPBS, cross 0.22 μm of filter membrane;
(3) preparation of colloid gold label thing: get the colloidal gold solution 40mL in step (1), use 0.25mol/LK
2cO
3regulate colloidal gold solution pH to 8.5, magnetic stirrer 250r/min stirs, and dropwise adds the protein solution of 4mL containing 0.3mg antibody protein, reaction 10min; Dropwise add 4mL10%BSA, continue stirring reaction 10min; The centrifugal 10min of gold labeling antibody solution normal temperature 1500r/min, discards the precipitation formed by the gold grain condensed; Red supernatant solution 4 DEG C, the centrifugal 20min of 12000r/min, abandons supernatant, collecting precipitation, and the golden labeling antibody dilution of precipitation is settled to 1mL, is prepared into carbendazim monoclonal antibody colloid gold label thing;
(4) preparation of colloidal gold film: step (3) carbendazim protein conjugate monoclonal antibody colloid gold label thing is sprayed on carrier glass cellulose membrane with drawing film instrument with the even concentration of 5 μ L/cm, room temperature is dried or 37 DEG C of oven dry 3h naturally, makes the colloidal gold film containing carbendazim protein conjugate monoclonal antibody colloid gold label thing;
(5) coated film preparation: sheep anti-mouse igg antibody, carbendazim protein conjugate are diluted to 1mg/mL, be sprayed on nitrocellulose filter with the concentration of 1 μ L/cm successively with drawing film instrument, be prepared into coated film, 37 DEG C of bags are by after 2h, and room temperature is dried or 37 DEG C of oven dry naturally;
(6) sample pad pre-treatment: be uniformly coated on by sample pad treating fluid on glass fibre element film, under the condition of air humidity lower than 60%, room temperature is dried naturally;
(7) assembling of test card: PVC offset plate pastes sample pad, colloidal gold film, coated film and absorbent wool from top to bottom successively, is assembled into test strips, cuts into the rectangular of one fixed width, then test strips is arranged in the test card shell of rectangular flat shelly.
2. carbendazim colloidal-gold detecting-card according to claim 1, is characterized in that the detection line T of institute's bag quilt in described step (5) is located at the below of nature controlling line C;
Sample pad treating fluid in described step (6) is by 1g Bovine serum albumin (BSA) and 0.8g sodium chloride (NaCl), is settled to 100mL with the 0.01mol/LPBS containing 0.5%TRITON-100.
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|---|---|---|---|
| CN201410551852.XA CN105572373A (en) | 2014-10-17 | 2014-10-17 | Immune colloidal gold detecting card of carbendazin and preparation method of immune colloidal gold detecting card |
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| CN201410551852.XA CN105572373A (en) | 2014-10-17 | 2014-10-17 | Immune colloidal gold detecting card of carbendazin and preparation method of immune colloidal gold detecting card |
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Cited By (14)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105424939A (en) * | 2015-12-23 | 2016-03-23 | 中国烟草总公司郑州烟草研究院 | Test paper strip for detecting carbendazim, preparation method and application thereof |
| CN106556694A (en) * | 2016-12-03 | 2017-04-05 | 百奥森（江苏）食品安全科技有限公司 | A kind of immune colloid gold detection card of carbendazim |
| CN108226500A (en) * | 2016-12-15 | 2018-06-29 | 江苏维赛科技生物发展有限公司 | The immune colloid gold detection blocking of fluoroquinolones Ciprofloxacin is standby |
| CN108226501A (en) * | 2016-12-15 | 2018-06-29 | 江苏维赛科技生物发展有限公司 | Immune colloid gold detection card of aflatoxin B1 and preparation method thereof |
| CN108226503A (en) * | 2016-12-15 | 2018-06-29 | 江苏维赛科技生物发展有限公司 | Immune colloid gold detection card of aflatoxin G 1 and preparation method thereof |
| CN108226502A (en) * | 2016-12-15 | 2018-06-29 | 江苏维赛科技生物发展有限公司 | Immune colloid gold detection card of Enrofloxacin and preparation method thereof |
| CN108226505A (en) * | 2016-12-15 | 2018-06-29 | 南京亿特生物科技有限公司 | Immune colloid gold detection card of sodium sulfocyanate and preparation method thereof |
| CN108226486A (en) * | 2016-12-15 | 2018-06-29 | 南京亿特生物科技有限公司 | Immune colloid gold detection card of Madumycin and preparation method thereof |
| CN109270263A (en) * | 2017-07-18 | 2019-01-25 | 中国医学科学院药用植物研究所 | The preparation and the application in Chinese medicine of a kind of carbendazim sxemiquantitative colloidal gold strip |
| CN109752533A (en) * | 2017-11-05 | 2019-05-14 | 江苏维赛科技生物发展有限公司 | A kind of immune colloid gold detection card and preparation method thereof detecting biphenyl |
| CN109752534A (en) * | 2017-11-05 | 2019-05-14 | 江苏维赛科技生物发展有限公司 | A kind of immune colloid gold detection card and preparation method thereof detecting o-phenyl phenol |
| CN109752544A (en) * | 2017-11-05 | 2019-05-14 | 江苏维赛科技生物发展有限公司 | A kind of colloidal gold strip detecting aflatoxin B1 |
| CN109752535A (en) * | 2017-11-05 | 2019-05-14 | 镇江华维检测技术有限公司 | The immune colloid of chloramphenicol detects card in a kind of detection tissue |
| 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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Cited By (15)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105424939A (en) * | 2015-12-23 | 2016-03-23 | 中国烟草总公司郑州烟草研究院 | Test paper strip for detecting carbendazim, preparation method and application thereof |
| CN105424939B (en) * | 2015-12-23 | 2017-06-20 | 中国烟草总公司郑州烟草研究院 | A kind of test strips for detecting carbendazim and its preparation method and application |
| CN106556694A (en) * | 2016-12-03 | 2017-04-05 | 百奥森（江苏）食品安全科技有限公司 | A kind of immune colloid gold detection card of carbendazim |
| CN108226502A (en) * | 2016-12-15 | 2018-06-29 | 江苏维赛科技生物发展有限公司 | Immune colloid gold detection card of Enrofloxacin and preparation method thereof |
| CN108226501A (en) * | 2016-12-15 | 2018-06-29 | 江苏维赛科技生物发展有限公司 | Immune colloid gold detection card of aflatoxin B1 and preparation method thereof |
| CN108226503A (en) * | 2016-12-15 | 2018-06-29 | 江苏维赛科技生物发展有限公司 | Immune colloid gold detection card of aflatoxin G 1 and preparation method thereof |
| CN108226500A (en) * | 2016-12-15 | 2018-06-29 | 江苏维赛科技生物发展有限公司 | The immune colloid gold detection blocking of fluoroquinolones Ciprofloxacin is standby |
| CN108226505A (en) * | 2016-12-15 | 2018-06-29 | 南京亿特生物科技有限公司 | Immune colloid gold detection card of sodium sulfocyanate and preparation method thereof |
| CN108226486A (en) * | 2016-12-15 | 2018-06-29 | 南京亿特生物科技有限公司 | Immune colloid gold detection card of Madumycin and preparation method thereof |
| CN109270263A (en) * | 2017-07-18 | 2019-01-25 | 中国医学科学院药用植物研究所 | The preparation and the application in Chinese medicine of a kind of carbendazim sxemiquantitative colloidal gold strip |
| CN109752533A (en) * | 2017-11-05 | 2019-05-14 | 江苏维赛科技生物发展有限公司 | A kind of immune colloid gold detection card and preparation method thereof detecting biphenyl |
| CN109752534A (en) * | 2017-11-05 | 2019-05-14 | 江苏维赛科技生物发展有限公司 | A kind of immune colloid gold detection card and preparation method thereof detecting o-phenyl phenol |
| CN109752544A (en) * | 2017-11-05 | 2019-05-14 | 江苏维赛科技生物发展有限公司 | A kind of colloidal gold strip detecting aflatoxin B1 |
| CN109752535A (en) * | 2017-11-05 | 2019-05-14 | 镇江华维检测技术有限公司 | The immune colloid of chloramphenicol detects card in a kind of detection tissue |
| 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: 20160511 |