CN101078727A - Colloidal gold silver combined test paper for antibody of canine animal and cat animal disease, and the preparation technology - Google Patents
Colloidal gold silver combined test paper for antibody of canine animal and cat animal disease, and the preparation technology Download PDFInfo
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Abstract
The invention provides an antibody colloid gold-silver combined detecting test paper for important blight of canidae and felid. Colloid-gold marker sheep-anti dog and cat IgG is mixed and is covered on glass fiber film to fabricate combination pad. Rarefied rabies virus, canine distemper virus, feline panleukopenia virus and canine parvovirus so on are covered at measuring lines R, D, F and P so on of nitrocellulose film. Sheep-anti-rabit IgG is covered at the quality control line C of nitrocellulose film. The detecting film is fabricated. When detected sample contains corresponding virus antibodies, it can be combined with gold marker sheep-anti dog and cat IgG firstly and then it is combined with corresponding antigen on the detecting line to form collection under the chromatography effect. And again it is colored strongly by silver dyeing solution to form color band visible by naked eyes and furthermore according to color result determination is carried out. The invention does not need help from other instrument equipment and measure multiple antibodies of important blight of canidae and felid at the same time. It is especially suitable for field detection and epidemiological investigation.
Description
Technical field:
The present invention relates to a kind of dog, the important eqpidemic disease antibody colloidal gold of cats Unionpay closes and detects test paper and technology of preparing thereof, be used for the antibody fast detecting of important eqpidemic diseases such as dog cats rabies, canine distemper, cat distemper heat and canine parvovirus disease, belong to animal epidemic detection technique field.
Background technology:
Dog, the important eqpidemic diseases of cats such as rabies, canine distemper, cat distemper heat and canine parvovirus disease be serious harm China fur economic animal industry not only, and the health of the uncommon wild animal of multiple treasure such as tiger, lion, wolf, the morbidity death that also can cause the people that has in serious threat.Will fundamentally control these eqpidemic diseases, most important means are immunity inoculations of vaccine, but whether have produced the antibody that reaches the immunoprotection level in the animal body after the vaccine inoculation, need to rely on certain technological means to detect.For dog, the important eqpidemic disease of cats, antibody detection method commonly used in the world at present has methods such as neutralization test, cell neutralization test, blood clotting-hemagglutination-inhibition test in immunofluorescence kitchen range inhibition test, enzyme linked immunosorbent assay, the mouse brain, but these methods all need certain instrument and equipment and higher operative technique, have influenced it and have applied.
Collaurum is a kind of detection technique that development in recent years is got up, at first be used for immuno-electron microscope by Faulk and Taylor (1971), has become additional greatly beyond fluorescein, radioactive isotope and the enzyme labeling now.Colloid gold test paper is to utilize antigen-antibody reaction and immunochromatography principle, with antigen or antibody with colloid gold label, when containing in the sample to be checked with the antibody of antigen or antibody specific bond or antigen, gathering will take place and produce macroscopic colour band, have easy, quick, visual result, advantage such as accurate.
Summary of the invention:
The invention provides a kind of dog, the detection test paper closes in the important eqpidemic disease antibody colloidal gold of cats Unionpay, can detect the antibody of the important eqpidemic diseases such as rabies, canine distemper, cat distemper heat and canine parvovirus disease that dog, feline body contain easy, quickly and accurately.
The invention also discloses the technology of preparing of above-mentioned detection test paper, be suitable for suitability for industrialized production.
Solution of the present invention be according to antigen-antibody can specific bond the immunology ultimate principle, utilize colloidal gold immunochromatographimethod technology and silver to dye the reinforcement technology, the goat-anti dog of colloid gold label and cat IgG are mixed being coated on the plain film of glass fibre, make pad; Detection line R, the D, F, the P etc. that hydrophobin (RV), CDV (CDV), feline distemper virus (FPV) and the canine parvovirus antigens such as (CPV) of purifying are coated on nitrocellulose filter (NC) respectively locate, the anti-sheep IgG of rabbit is coated on the nature controlling line C place of nitrocellulose filter, makes the detection film; When containing corresponding antiviral antibody in the test sample, can combine with gold mark goat-anti dog and cat IgG earlier, then under the chromatography effect with the respective detection line on antigen combine and enrichment, strengthen the dyeing back through silver-colored dye liquor again and form macroscopic colour band, and then judge according to the result that develops the color.
In addition, preparation method provided by the invention relates generally to the content of the following aspects:
1, viral antigen preparation: with rabies viruses (RV), CDV (CPV), feline distemper virus (FPV) and canine parvovirus (CPV) etc. inoculate respectively Vero cell, F81 cell etc. separately sensitive cells cultivate propagation; concentrate purification then respectively; purifying antigens such as preparation RV, CDV, FPV and CPV; with protective agent, standby in-30 ℃ of preservations.
2, goat-anti dog and cat IgG preparation: with purified dog and cat serum IgG difference immune sheep, preparation goat-anti dog and cat serum, and extract its IgG, preparation goat-anti dog and cat IgG, with protective agent, standby in-30 ℃ of preservations.
3, gold mark goat-anti dog and cat IgG preparation: carry out colloid gold label respectively with prepared goat-anti dog and cat IgG, preparation gold mark goat-anti dog and cat IgG, adding preservative agent, standby in 4 ℃ of preservations.
4, the anti-sheep IgG preparation of rabbit: with the sheep IgG immunizing rabbit of purifying, make the anti-sheep immune serum of rabbit, and therefrom extract IgG, the anti-sheep IgG of preparation rabbit, with protective agent, standby in-30 ℃ of preservations.
5, test paper preparation and assembling: press shown in the accompanying drawing, after earlier gold being marked goat-anti dog and cat IgG and being mixed by a certain percentage, bag is made pad by in glass fibre element film; Respectively the antigen speckings such as RV, CDV, FPV and CPV of purifying are located in detection line R, D, F, the P etc. of nitrocellulose filter (NC) then,, made the detection film the nature controlling line C place of the anti-sheep IgG of rabbit specking in the NC film; Down payment mark chromatographic test paper prepares requirement more at last, sample pad, pad, detection film and absorption pad are carried out adhesion, suppress and cut, make dog, the detection test paper closes in the important eqpidemic disease antibody colloidal gold of cats Unionpay, be loaded on and make test card in the plastic clip, add the drying agent packing, be closed in the polybag, in 4 ℃ of kept dry.
6, the preparation of silver-colored reinforcement corresponding reagent:
The preparation of activating solution: with the citrate buffer of deionized water preparation 0.05mol/L, PH3.8.
The preparation of silver dye liquor: silver acetate, p-dihydroxy-benzene, gum arabic are dissolved in the deionized water, make that its final concentration is respectively 0.11%, 0.35%-1.7% and 7%-10%.
The preparation of stop bath: 20% sodium thiosulfate solution.
During detection, test paper is lain against desktop, get serum to be checked or anticoagulated whole blood 50-100 microlitre drips in well, wash with activating solution after 10-15 minute, drip 30-50 microlitre silver dye liquor to detecting film again, with deionized water wash, Dropwise 5 0-100 microlitre stop bath was judged after 8-10 minute again after 5 minutes.Occur at nature controlling line under the prerequisite of obvious colour band, detection line the colour band occurs, and the person then judges its corresponding antibodies positive, otherwise then is judged to be negative antibody; Colour band do not occur as nature controlling line, show that detectable lost efficacy, retrial need ascertain the reason.
The present invention has following advantage compared with prior art: easy, suitable, quick, economic, pollution-free, the easy to operate and interpretation of the present invention, need be by Other Instruments equipment, and can detect the important eqpidemic disease antibody of multiple dog cats simultaneously, be specially adapted to on-the-spot detection and epidemiology survey etc.Preparation method's science of the present invention, cost is low, stability is high, good reproducibility, is easy to make, storage and transport.
Description of drawings
Fig. 1 is the structural representation that the present invention detects test paper:
Fig. 2 is that the present invention detects the test paper sectional view.
1-plastic clip; 2-well; 3-viewport; 4-sample pad; 5-pad (make by the plain film of glass fibre, be coated with gold mark goat-anti dog and cat IgG); 6-detection film (made by nitrocellulose filter, specking has several detection lines and 1 nature controlling line on it); 7-absorption pad; 8-liner plate.R, D, F, P etc. are detection line, are coated with purifying antigens such as RV, CDV, FPV and CPV respectively; C is a nature controlling line, is coated with the anti-sheep IgG of rabbit.
Embodiment
By following examples the present invention is described for example further, and do not limit the present invention in any way, under the prerequisite that does not deviate from technical solution of the present invention, any change or change that those of ordinary skills that the present invention did are realized easily all will fall within the claim scope of the present invention.
Embodiment 1
According to shown in Figure 1, it is as follows that the present invention detects the test paper structure:
1, viral antigen preparation: with rabies viruses (RV), CDV (CPV), feline distemper virus (FPV) and canine parvovirus (CPV) etc. inoculate respectively Vero cell, F81 cell etc. separately sensitive cells cultivate propagation; concentrate purification then respectively; purifying antigens such as preparation RV, CDV, FPV and CPV; with protective agent, standby in-30 ℃ of preservations.
2, goat-anti dog and cat IgG preparation: with purified dog and cat serum IgG difference immune sheep, preparation goat-anti dog and cat serum, and extract its IgG, preparation goat-anti dog and cat IgG, with protective agent, standby in-30 ℃ of preservations.
3, gold mark goat-anti dog and cat IgG preparation: carry out colloid gold label respectively with prepared goat-anti dog and cat IgG, preparation gold mark goat-anti dog and cat IgG, adding preservative agent, standby in 4 ℃ of preservations.
4, the anti-sheep IgG preparation of rabbit: with the sheep IgG immunizing rabbit of purifying, make the anti-sheep immune serum of rabbit, and therefrom extract IgG, the anti-sheep IgG of preparation rabbit, with protective agent, standby in-30 ℃ of preservations.
5, test paper preparation and assembling: press shown in the accompanying drawing, after earlier gold being marked goat-anti dog and cat IgG and being mixed by a certain percentage, bag is made pad by in glass fibre element film; Respectively the antigen speckings such as RV, CDV, FPV and CPV of purifying are located in detection line R, D, F, the P etc. of nitrocellulose filter (NC) then,, made the detection film the nature controlling line C place of the anti-sheep IgG of rabbit specking in the NC film; Down payment mark chromatographic test paper prepares requirement more at last, sample pad, pad, detection film and absorption pad are carried out adhesion, suppress and cut, make dog, the detection test paper closes in the important eqpidemic disease antibody colloidal gold of cats Unionpay, be loaded on and make test card in the plastic clip, add the drying agent packing, be closed in the polybag, in 4 ℃ of kept dry.
6, the preparation of silver-colored reinforcement corresponding reagent:
The preparation of activating solution: with the citrate buffer of deionized water preparation 0.05mol/L, PH3.8.
The preparation of silver dye liquor: silver acetate, p-dihydroxy-benzene, gum arabic are dissolved in the deionized water, make that its final concentration is respectively 0.11%, 0.35%-1.7% and 7%-10%.
The preparation of stop bath: 20% sodium thiosulfate solution.
Claims (2)
1, the detection test paper closes in a kind of dog, the important eqpidemic disease antibody colloidal gold of cats Unionpay, it is characterized in that: liner plate 8 big envelopes are in plastic clip 1, the surface of plastic clip 1 offers well 2 and viewport 3, is assembled with treated sample pad 4, pad 5 on the surface of liner plate 8, detects film 6 and absorption pad 7.Wherein, pad 5 is made by the plain film of glass fibre, on be coated with the goat-anti dog and the cat IgG of colloid gold label; Detect film 6 and made by nitrocellulose filter, last specking has many detection lines and 1 nature controlling line, and R, D, F, P etc. are detection line, wrap respectively by purifying antigens such as RV, CDV, FPV and CPV, and C is a nature controlling line, is coated with the anti-sheep IgG of rabbit; Detecting film links to each other with absorption pad 7.
2, detection method for preparing test paper according to claim 1 comprises the steps:
1) viral antigen preparation: with rabies viruses (RV), CDV (CPV), feline distemper virus (FPV) and canine parvovirus (CPV) etc. inoculate respectively Vero cell, F81 cell etc. separately sensitive cells cultivate propagation, concentrate respectively then and purify, purifying antigens such as preparation RV, CDV, FPV and CPV, with protective agent, standby in-30 ℃ of preservations;
2) goat-anti dog and cat IgG preparation: with purified dog and cat serum IgG difference immune sheep, preparation goat-anti dog and cat serum, and extract its IgG respectively, preparation goat-anti dog and cat IgG, with protective agent, standby in-30 ℃ of preservations;
3) gold mark goat-anti dog and cat IgG preparation: carry out colloid gold label respectively with prepared goat-anti dog and cat IgG, preparation gold mark goat-anti dog and cat IgG, adding preservative agent, standby in 4 ℃ of preservations;
4) the anti-sheep IgG preparation of rabbit: with the sheep IgG immunizing rabbit of purifying, make the anti-sheep immune serum of rabbit, and therefrom extract IgG, the anti-sheep IgG of preparation rabbit, with protective agent, standby in-30 ℃ of preservations;
5) test paper preparation and assembling: press shown in the accompanying drawing, after earlier gold being marked goat-anti dog and cat IgG and being mixed by a certain percentage, bag is made pad by in glass fibre element film; Respectively the antigen speckings such as RV, CDV, FPV and CPV of purifying are located in detection line R, D, F, the P etc. of nitrocellulose filter (NC) then,, made the detection film the nature controlling line C place of the anti-sheep IgG of rabbit specking in the NC film; Down payment mark chromatographic test paper prepares requirement more at last, sample pad, pad, detection film and absorption pad are carried out adhesion, suppress and cut, make dog, the detection test paper closes in the important eqpidemic disease antibody colloidal gold of cats Unionpay, be loaded on and make test card in the plastic clip, add the drying agent packing, be closed in the polybag, in 4 ℃ of kept dry;
6) preparation of silver-colored reinforcement corresponding reagent:
The preparation of activating solution: with the citrate buffer of deionized water preparation 0.05mol/L, PH3.8;
The preparation of silver dye liquor: silver acetate, p-dihydroxy-benzene, gum arabic are dissolved in the deionized water, make that its final concentration is respectively 0.11%, 0.35%-1.7% and 7%-10%;
The preparation of stop bath: 20% sodium thiosulfate solution.
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| Application Number | Priority Date | Filing Date | Title |
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| CN 200710055760 CN101078727A (en) | 2007-06-13 | 2007-06-13 | Colloidal gold silver combined test paper for antibody of canine animal and cat animal disease, and the preparation technology |
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| CN 200710055760 CN101078727A (en) | 2007-06-13 | 2007-06-13 | Colloidal gold silver combined test paper for antibody of canine animal and cat animal disease, and the preparation technology |
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Cited By (11)
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| CN102707061A (en) * | 2012-06-18 | 2012-10-03 | 深圳市博卡生物技术有限公司 | Gold immunochromatographic assay quantitative detection test paper and detection method |
| CN101738470B (en) * | 2008-11-06 | 2013-04-03 | 益思美诠生物科技(上海)有限公司 | Immunochromatography method capable of simultaneously detecting various analytes |
| CN103529222A (en) * | 2013-10-29 | 2014-01-22 | 华中农业大学 | Rabies virus antibody colloidal gold detection test strip for human and livestock joint detection and preparation method thereof |
| CN103743901A (en) * | 2013-12-19 | 2014-04-23 | 河北北方学院 | Colloidal gold test strip for detecting canine parvovirus |
| CN104360061A (en) * | 2014-11-20 | 2015-02-18 | 长春西诺生物科技有限公司 | Rabies virus IgG antibody immune gold-labeled test paper and preparation method thereof |
| CN105579846A (en) * | 2013-09-12 | 2016-05-11 | 阿瑟斯生物有限公司 | Apparatus for manufacturing diagnosis kit, and diagnosis kit manufactured by same |
| CN105891491A (en) * | 2016-04-11 | 2016-08-24 | 洛阳普莱柯万泰生物技术有限公司 | Kit and application thereof |
| CN109813904A (en) * | 2019-03-27 | 2019-05-28 | 丁建红 | African swine fever virus rapid detection card |
| CN110302374A (en) * | 2019-08-09 | 2019-10-08 | 中国农业科学院特产研究所 | Canine vaccine and its preparation method and application |
| CN110568204A (en) * | 2019-06-10 | 2019-12-13 | 青岛海润检测股份有限公司 | Chemiluminescence kit for one-step quantitative detection of canine distemper virus-canine parvovirus antibody |
| CN111505279A (en) * | 2020-04-17 | 2020-08-07 | 南昌大学 | Silver growth-enhanced ultra-sensitive rotavirus detection method |
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Cited By (14)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101738470B (en) * | 2008-11-06 | 2013-04-03 | 益思美诠生物科技(上海)有限公司 | Immunochromatography method capable of simultaneously detecting various analytes |
| CN102707061A (en) * | 2012-06-18 | 2012-10-03 | 深圳市博卡生物技术有限公司 | Gold immunochromatographic assay quantitative detection test paper and detection method |
| CN105579846B (en) * | 2013-09-12 | 2018-06-05 | 阿瑟斯生物有限公司 | For manufacturing the diagnostic kit of the device of diagnostic kit and its manufacture |
| CN105579846A (en) * | 2013-09-12 | 2016-05-11 | 阿瑟斯生物有限公司 | Apparatus for manufacturing diagnosis kit, and diagnosis kit manufactured by same |
| CN103529222A (en) * | 2013-10-29 | 2014-01-22 | 华中农业大学 | Rabies virus antibody colloidal gold detection test strip for human and livestock joint detection and preparation method thereof |
| CN103743901A (en) * | 2013-12-19 | 2014-04-23 | 河北北方学院 | Colloidal gold test strip for detecting canine parvovirus |
| CN104360061A (en) * | 2014-11-20 | 2015-02-18 | 长春西诺生物科技有限公司 | Rabies virus IgG antibody immune gold-labeled test paper and preparation method thereof |
| CN105891491A (en) * | 2016-04-11 | 2016-08-24 | 洛阳普莱柯万泰生物技术有限公司 | Kit and application thereof |
| CN109813904A (en) * | 2019-03-27 | 2019-05-28 | 丁建红 | African swine fever virus rapid detection card |
| CN110568204A (en) * | 2019-06-10 | 2019-12-13 | 青岛海润检测股份有限公司 | Chemiluminescence kit for one-step quantitative detection of canine distemper virus-canine parvovirus antibody |
| CN110568204B (en) * | 2019-06-10 | 2022-04-01 | 青岛海润检测股份有限公司 | Chemiluminescence kit for one-step quantitative detection of canine distemper virus-canine parvovirus antibody |
| CN110302374A (en) * | 2019-08-09 | 2019-10-08 | 中国农业科学院特产研究所 | Canine vaccine and its preparation method and application |
| CN110302374B (en) * | 2019-08-09 | 2023-04-14 | 黑龙江八一农垦大学 | Canine vaccine and preparation method and application thereof |
| CN111505279A (en) * | 2020-04-17 | 2020-08-07 | 南昌大学 | Silver growth-enhanced ultra-sensitive rotavirus detection method |
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