CN105277702A - Preparation of colloidal gold immunochromatographic detection card for detecting O,O-dimethyl-O-2,2-dichlorovinylphosphate and detection method - Google Patents
Preparation of colloidal gold immunochromatographic detection card for detecting O,O-dimethyl-O-2,2-dichlorovinylphosphate and detection method Download PDFInfo
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- CN105277702A CN105277702A CN201410350262.0A CN201410350262A CN105277702A CN 105277702 A CN105277702 A CN 105277702A CN 201410350262 A CN201410350262 A CN 201410350262A CN 105277702 A CN105277702 A CN 105277702A
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- detection
- ddvp
- dimethyl
- dichlorovinylphosphate
- colloidal gold
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Abstract
The invention discloses an O,O-dimethyl-O-2,2-dichlorovinylphosphate colloidal gold detection card, and is mainly used for rapid detection of the content of O,O-dimethyl-O-2,2-dichlorovinylphosphate in food. According to a method, a sample liquid absorbing part, a colloidal gold marking part, a detection reaction part and a water absorbing part are orderly pasted on a backing of test paper. The detection reaction part is coated by one bar of an antigen which is applied for detection and is used as a detection line, and at the same time, the detection reaction part is also coated by one bar of an IgG which resists the second species of animal protein and is used as a reference line. The rapid detection test paper strip has high specificity, can carry out semi quantitative detection, can be used at the environmental temperature of 4 to 35 DEG C, and is suitable for an individual farmer, a food hygiene and quality inspection department, customs and other animal-derived food for rapid detection on O,O-dimethyl-O-2,2-dichlorovinylphosphate residues. The colloidal gold immunochromatographic detection card has the beneficial effects of high specificity, high sensitivity, simple and convenient operation and the like, and can carry out semi-quantitative detection.
Description
Technical field
Belong to field of detection of food safety, be specifically related to the detection method of the Harmful Residue in food, particularly detect the preparation of the colloidal gold immunochromatographimethod test card of DDVP.
Background technology
DDVP (English name O, O-dimethyl-O-2,2-dichlorovinylphosphate) has another name called DDVP, formal name used at school O, O-dimethyl-O-(2,2-dichloroethylene) phosphate, and organic phosphorous insecticide is a kind of, molecular formula C4H7Cl2O4P.A kind of organophosphorus insecticide, industrial products are colourless to light brown liquid, and sterling boiling point 74 DEG C (under 133.322Pa) volatility is large, under room temperature in water solubleness 1%, solubleness 2% ~ 3% in kerosene, can be dissolved in organic solvent, facile hydrolysis, chance caustic digestion is faster.Toxicity is large.Colourless oil liquid, has volatility, is used for the agricultural pests such as prevention and control of aphides, is also used for killing mosquito, fly etc.Available missible oil be made into high concentration medicine liquid spray stored grain insects, evil mite, ventilate after airtight 3 ~ 4 days loose gas again, when temperature is higher, drug effect is better.Medicine liquid spray is stifling on the metope of cotton storehouse, can water prevention brown plant-hopper, pink bollworm.Pesticide-clay mixture or malicious chaff field are spread fertilizer over the fields stifling, can prevent and treat mythimna separata.Medicine liquid spray, can prevent and treat the hiding pests such as rice leaf roller.DDVP can decompose rapidly after using, lasting period is short, noresidue, can use in period very short before crop harvesting, to prevent and treat sucking mouth parts and pests with chewing mouthparts, therefore being applicable on the fruit trees such as apple, pears, grape and vegetables, mushroom, tea tree, mulberry tree, tobacco, before general results, disable period is about 7 days.Easily poisoning is occurred to jowar, corn, melon, beans is also more responsive, should note during use.China GB5127-85 specifies the permitting deformation 0.2mg/kg(veterinary antibiotics of organophosphorus pesticide in food).
Summary of the invention
The object of the invention is to provide and detects the residual of DDVP in food fast.
Technical scheme of the present invention is the test card of DDVP and the disposal route of detection sample thereof.The test card of DDVP arranges detector bar in the test card shell of rectangular flat shell shape, test card case surface has detection fenestra and well, it is characterized in that test-strips forms by four kinds of original papers pasted by supporting backboard: in the middle part of supporting backboard, paste nitrocellulose filter, thieving paper is pasted in backboard one end in supporting, the other end pastes sample pad, thieving paper is inner to be overlapped with nitrocellulose filter, then there is one section of gold mark pad between sample pad and nitrocellulose filter, by gold mark pad, sample pad and nitrocellulose filter are linked together.It is the potpourri containing the second kind albumen and DDVP antibody in gold mark pad, nitrocellulose filter laterally there are 2 isolated colour developing marking bands, article one, be the detection zone containing DDVP protein conjugate, another be the IgG(of the second kind animal protein conventional be anti-rabbit antibody or anti-mouse antibody); Test-strips is inserted in test card shell, and sample pad is just to well, and nitrocellulose filter is just to detection fenestra.
Sample treatment is: take sample 2 ± 0.05g in the centrifuge tube of 50mL, add 10mL ultrapure water, smash (about 1min) with high speed tissue pulverizer under 1800r/min condition, get viscous fluid and add 50% trichloroacetic acid 2mL and methylene chloride 10mL, high speed jolting 30min on Clothoid type oscillator.Then centrifugal 30min under 4 DEG C of 5000rpm conditions, gets supernatant 10mL, and add methylene chloride 2mL, on cyclotron vibration mixing 20min, then under 4 DEG C of 12000rpm conditions centrifugal 30min, supernatant is the extract of medicine.
Advantage of the present invention can detect the DDVP contained in sample fast, and save testing cost, easy to use, and detect fast, highly sensitive, result is accurate.
Accompanying drawing explanation
Fig. 1 is DDVP test card shell and test strip arrangement figure in the enclosure.
Fig. 2 is DDVP test card test strip structural representation.
Fig. 3 test strip nitrocellulose filter shows trace band schematic diagram.
Embodiment
1. mark the preparation of collaurum
The preparation of colloidal gold solution: first by 1g gold chloride (HAuCl
4) powder is dissolved in the chlorauric acid solution that 100mL ultrapure water is configured to 1%; Get 1L ultrapure water again to heat on speciality electric furnace, the trisodium citrate aqueous solution of 10mL1% is added to boiling, heating is continued after boiling, the chlorauric acid solution of 10mL1% is added after 5 minutes, again be heated to boiling, wait for that color starts timing after becoming claret, after 10 minutes, stop heating, take off and naturally to cool afterwards, be settled to 1L, be colloidal gold solution.
Antibody labeling: determine the 0.1mol/lK needed for antibody
2cO
3ratio, get 100mL colloidal gold solution, add the K of this ratio 0.1mol/L
2cO
3mixing (about 5min), again DDVP and the second kind animal protein are used PBS(0.01mo1/L respectively, pH7.4) dissolved dilution is to 2mg/mL, add the DDVP antibody (or 2mL2mg/mL second kind animal protein) of 2mL2mg/mL, mixing (5min), finally adds 10%BSA2mL, mixing (5min).Stay precipitation centrifugal three times, the precipitation of collecting for these three times is gold mark DDVP antibody.
Colloid gold label part process: after DDVP antibody dilution, soaks into carrier (as glass fibre, polyester film etc.) by a certain percentage, and evenly, taking-up is put in 37 DEG C of constant temperature ovens and smokes 8-12h, namely as colloid gold label part after drying paving.
2. the special polymer compound film preparation of immunochromatography
First make transparent backing film with a kind of nitrocellulose filter, backing film covers one deck strong to protein bound power, the chromatographic film of certain pore size that what wetting property was good have.The effect of backing film stops organic solvent in bonding agent to the destruction of protein in chromatographic film.
3. the spotting methods of chromatographic film
By certain density happy antigen (conjugates that DDVP and carrier mass are formed) accurate quantification; Schedule in chromatographic film in horizontal stripe shape point, for detection with a determining deviation (5mm).
DDVP test card arranges test strip 2 in test card shell 1, structure and the arranging of test strip 2 of test card shell 1 see accompanying drawing 1, test card shell 1 is rectangular flat shell shape shell, long 70mm, wide 20mm, thick 5mm, shell wall thickness 1mm, be made up of engineering plastics, test card shell 1 is formed by upper cover and lower cover two semi join, test card shell 1 has covered and has detected fenestra 3 and well 4.Test strip 2 is placed in the lower cover of test card shell 1.
Test strip 2 is fillet thin slices of sandwich construction, length of a film 60mm, and the wide about 4mm of sheet, thickness is less than 2.5mm.Test strip 2 is sandwich constructions: bottom is supporting backboard 5, and be polyethylene sheets, thickness is about 0.5mm, long 60mm, wide 4mm; Nitrocellulose filter 6 is pasted, cellulose nitrate thickness 0.5mm, long 20mm, wide 4mm in the middle part of supporting backboard 5.Nitrocellulose filter 6 has two isolated laterally display trace bands, see accompanying drawing 3, in two display traces, one is detection line 10, and containing DDVP conjugate, another is line of reference 11, containing the IgG of the second kind animal protein.Thieving paper 9 is pasted in backboard 5 one end in supporting, the other end pastes sample pad 8, and thieving paper 9 is inner to be overlapped with nitrocellulose filter 6, then has one section of gold to mark pad 7 between sample pad 8 and nitrocellulose filter 6, sample pad 8 and nitrocellulose filter 6 are linked together by gold mark pad 7, lap width is at 1-2mm.Sample pad 8 material is water adsorption glass fiber or polyester film.
When using DDVP colloidal-gold detecting-card, first by sample liquid to be detected (after the extract dilution of medicine, be directly used in detection) from well 4, instill the sample pad 8 of DDVP test strip 2, due to rainbow action principle, the dichlorvos monoclonal antibody colloid gold label thing against the enemy contained by sample liquid to be detected and colloidal gold film 7 is driven to spread to nitrocellulose filter 6 together, observations in 5-10 minute.
The key reaction of test card is immunologic antigen and antibody response, the antibody of the colloid gold label that nitrocellulose filter moves, on p-wire with containing DDVP protein conjugate, and line of reference reacts containing the IgG of the second kind animal protein, formation brownish red band.If corresponding DDVP to be measured is higher than permissible value in sample, sample adds rear elder generation and marks the antibody response in padding with gold, and can not with detection zone with DDVP protein conjugate react, thus not develop the color.Mainly contain following 3 kinds of testing results:
1, detection line 10 and line of reference 11 manifest brownish red trace simultaneously, represent that testing result be negative, illustrate in sample and do not contain DDVP or content lower than permissible value.
2, detection line 10 does not develop the color, and line of reference 11 manifests brownish red trace, represents that testing result is positive, illustrates that in sample, DDVP content is higher than permissible value.
3, detection line 10 manifests brownish red trace, and line of reference 11 does not develop the color, or detection line 10 and line of reference 11 all do not develop the color, and represents that test strip lost efficacy.
Claims (4)
1. DDVP colloidal-gold detecting-card, it is characterized in that: on the backing of test paper, post sample liquid absorption portion, colloid gold label part, detection reaction part and water absorbent portion successively, the material that colloid gold label part is labeled is the potpourri of the second kind animal protein and DDVP antibody; Detection reaction part is coated with detection DDVP antigen 1 bar as detection line, is also coated with the IgG1 bar of anti-second kind animal protein as line of reference simultaneously.
2. DDVP colloidal-gold detecting-card according to claim 1, is characterized in that: DDVP detection antigen is the conjugates that DDVP and carrier mass are formed.
3. DDVP colloidal-gold detecting-card according to claim 2, is characterized in that: described carrier mass, is protein, protein fragments, improvement on synthesis, semi-synthetic polypeptide or polysaccharide.
4. DDVP colloidal-gold detecting-card according to claim 1, is characterized in that: the second described kind animal protein is the protein of non-antibody source animal.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410350262.0A CN105277702A (en) | 2014-07-23 | 2014-07-23 | Preparation of colloidal gold immunochromatographic detection card for detecting O,O-dimethyl-O-2,2-dichlorovinylphosphate and detection method |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410350262.0A CN105277702A (en) | 2014-07-23 | 2014-07-23 | Preparation of colloidal gold immunochromatographic detection card for detecting O,O-dimethyl-O-2,2-dichlorovinylphosphate and detection method |
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| CN105277702A true CN105277702A (en) | 2016-01-27 |
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101042405A (en) * | 2007-04-26 | 2007-09-26 | 浙江省农业科学院 | Immune colloidal gold test strip for detecting organophosphorus pesticide and method for making same |
| CN102520165A (en) * | 2011-12-29 | 2012-06-27 | 北京康美天鸿生物科技有限公司 | Method for highly sensitive quantitative detection of quantum dot fluorescence immunochromatographic assay |
| CN103698510A (en) * | 2012-12-21 | 2014-04-02 | 欧普图斯(苏州)光学纳米科技有限公司 | Analyzing chemical and biological substances using nano-structure based spectral sensing |
| CN103808939A (en) * | 2012-11-06 | 2014-05-21 | 江苏维赛科技生物发展有限公司 | Spectinomycin colloidal gold detection card |
-
2014
- 2014-07-23 CN CN201410350262.0A patent/CN105277702A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101042405A (en) * | 2007-04-26 | 2007-09-26 | 浙江省农业科学院 | Immune colloidal gold test strip for detecting organophosphorus pesticide and method for making same |
| CN102520165A (en) * | 2011-12-29 | 2012-06-27 | 北京康美天鸿生物科技有限公司 | Method for highly sensitive quantitative detection of quantum dot fluorescence immunochromatographic assay |
| CN103808939A (en) * | 2012-11-06 | 2014-05-21 | 江苏维赛科技生物发展有限公司 | Spectinomycin colloidal gold detection card |
| CN103698510A (en) * | 2012-12-21 | 2014-04-02 | 欧普图斯(苏州)光学纳米科技有限公司 | Analyzing chemical and biological substances using nano-structure based spectral sensing |
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Application publication date: 20160127 |