CN1975428A - Sword-leaved cymbidium leaf virus spot enzyme-linked assay kit, preparation thereof and detecting method - Google Patents
Sword-leaved cymbidium leaf virus spot enzyme-linked assay kit, preparation thereof and detecting method Download PDFInfo
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Abstract
The invention discloses double-antibody sandwich ELSA regent box and the method for the Cymbidiummosaicvirus spot. The box is made up of the box body, every using liquid and the PVDF film. The preparation of the regent box is simple, quick, safe and reliable. The sample can be used little and it can be detected in a little time. Comparing to other serology detecting method, the method is simple, quick, sensitivity and high specialty, also it does not need the special device and get the result by the eye directly. So it can be used widely in the base unit.
Description
Technical field
The present invention relates to detect kit and the preparation and the detection method of cymbidium mosaic virus, particularly relate to sword-leaved cymbidium leaf virus spot enzyme-linked assay kit and prepare the method for this kit and the detection method that this kit of application carries out the dot enzyme-linked detection of cymbidium mosaic virus.
Background technology
Cymbidium mosaic virus (Cymbidium mosaic virus, CyMV) be subordinate to potexvirus, can make sword-leaved cymbidium produce chlorisis spot and necrotic plaque, Bowring cattleya produces local necrosis, tree orchid belongs to the generation floral leaf, in addition, also infecting tropical spice crop vanilla, is one of various main viruses of viewing and admiring orchid of harm.In view of the harmfulness of CyMV to China's orchid industry, it is very important to study its detection technique.
Plant virus detects enzyme linked immunosorbent assay (ELISA) (ELISA) method commonly used, EIA enzyme immunoassay is the technology that the specificity with the antigen-antibody combination combines with the efficient catalytic of enzyme, have highly sensitive, characteristics such as high specificity, but operation steps is many, length consuming time, and need relatively costly enzyme labelled antibody and 96 hole ELISA Plate.
Dot enzyme-linked detection is that the principle with the enzyme immuning adsorpting analysis is the detection method that the basis develops rapidly in recent years, once this method of research report was used for the salmonella and the conventional separation and Culture detection comparison of fast detecting sheep and goat carcass, not only easy, quick, and safety, reliable.And not relevant as yet at present spot immune enzyme coupling is in bibliographical information that detects cymbidium mosaic virus and the dot enzyme-linked detection kit product that is used to detect this virus.
Summary of the invention
The object of the present invention is to provide the kit of easy, quick, safe, the reliable and economical and practical dot enzyme-linked detection of cymbidium mosaic virus.
Another object of the present invention is to provide the method for preparing sword-leaved cymbidium leaf virus spot enzyme-linked assay kit.
A further object of the present invention is to provide the sword-leaved cymbidium leaf virus spot enzyme-linked assay kit that utilizes preparation to carry out the detection method of dot enzyme-linked detection cymbidium mosaic virus.
For achieving the above object, technical solution of the present invention is:
Sword-leaved cymbidium leaf virus spot enzyme-linked assay kit, mainly by box body, various bottled liquid and the pvdf membrane used that is located in the box body formed.
Described various bottledly comprise with liquid:
1 bottle of insulation liquid I
1 bottle of insulation liquid II
1 bottle of colour developing damping fluid
1 of developer
1 of positive
1 of negative sample
1 bottle of sample preparation liquid
1 bottle of lavation buffer solution
Described insulation liquid I is the 0.01-0.1M phosphate buffer that contains the cymbidium mosaic virus polyclonal antibody, includes 1% gelatin.
Described insulation liquid II is the 0.01-0.1M phosphate buffer that contains goat-anti rabbit enzyme labelled antibody, includes 1% gelatin.
Described colour developing damping fluid is 0.05-0.3M, and pH9.5 contains MgCl
2The Tris damping fluid of 10g/L+NaCl2g/L.
Described developer is 0.01g/ml NBT+0.005g/ml BCIP.
Described positive is the cymbidium mosaic virus of purifying.
Described positive is the cymbidium mosaic virus of purifying, and concentration is 0.1 μ g-100g/ml scope.
Described negative sample grinds the solution of healthy leaf preparation for sealing buffer solution.
Described sample preparation liquid is 1-5M, the carbonate buffer solution of pH9.6.
Described lavation buffer solution is 0-0.05M, the phosphate buffer of pH7.4.
The preparation method of described sword-leaved cymbidium leaf virus spot enzyme-linked assay kit, it mainly comprises the following steps:
1, the extraction of cymbidium mosaic virus and purifying: after getting the homogenate of the sick leaf adding of 100g datura 100ml0.5M citrate buffer solution (pH6.5), add the 100ml chloroform and stirred 30 minutes.At Beckman 10, centrifugal 20 minutes of 000rpm (JA-10).Get the NaCl that supernatant adds 4%PEG6000 and 0.3mol, left standstill after the stirring 1 hour.At Beckman 10, centrifugal 20 minutes of 000rpm (JA-10) abandons supernatant, fully repeats the PEG precipitation one time behind the dissolution precipitation with 0.5M citrate buffer solution (pH6.5).After getting for the second time the suspension of post precipitation usefulness 5mM borate buffer, at Beckman 10, centrifugal 20 minutes of 000rpm (JA-10) gets supernatant and uses P42A rotary head 40, centrifugal 2 hours of 000rpm in ultracentrifuge.Get precipitation and suspend again, carry out differential centrifugation again one time, get precipitation at last and dissolve with an amount of 0.01MPB (pH7.0) with 0.01MPB (pH7.0).If impurity is too many, carry out differential centrifugation again one time.Get thick purified virus and place 10%-40% sucrose pad gradient, use P42A rotary head 40, centrifugal 2 hours of 000rpm in ultracentrifuge.Collect viral band, use P42A rotary head 40 in ultracentrifuge once more, centrifugal 2 hours precipitation virus of 000rpm is dissolved among the 0.01MPB (pH7.0) at last, place-40 ℃ standby.
2, sero-fast preparation and purifying: select for use male rabbit to do immune animal, the purified virus that adds the emulsification of equivalent FreundShi Freund is done immunogene, adopts three intramuscular injection and twice intravenous immune rabbit.The amount of per injection virus is respectively 0.5mg, 0.75mg, 1mg, 1.5mg and 2mg, and be 7 days interval time, and last injection back blood sampling in 7-10 days 2 times is separated out serum and tired with agar double immunodiffusion method mensuration; By the sad precipitation method in conjunction with DEAE ion-exchange chromatography antibody purification.Regulate antiserums to pH4.8 with 2 times of volume 0.1M ammonium acetates, press 0.75ml sad/1ml serum adds caprylic acid, mixes and stirs 1 hour, centrifugal 30 minutes of 5000rpm gets supernatant, places 10mM Tris-Cl pH8.5 damping fluid to spend the night bag filter, the desalination of dialysing.Get and dialyse completely that solution is splined in the ion exchange column, with 10mMTris-HCl pH8.5 damping fluid 5ml/min flushing 15 minutes, carry out wash-out with the 10mM Tris-HCl pH8.5 buffer concentration gradient that contains 0-1M NaCl again, collect the eluting peak part, concentrate dialysis, obtain the cymbidium mosaic virus polyclonal antibody of purifying.
3, the preparation of the positive, negative sample:
Positive is the cymbidium mosaic virus virus of purifying, and concentration is in 0.1 μ g-100 μ g/ml scope; Negative sample grinds the solution of healthy leaf preparation for sealing buffer solution.
4, various preparations with liquid:
Insulation liquid I is the 0.01-0.1M phosphate buffer that contains the cymbidium mosaic virus polyclonal antibody, includes 1% gelatin.
Insulation liquid II is the 0.01-0.1M phosphate buffer that contains goat-anti rabbit enzyme labelled antibody, includes 1% gelatin.
The colour developing damping fluid is 0.05-0.3M, and pH9.5 contains MgCl
2The Tris damping fluid of 10g/L+NaCl 2g/L.
Developer is 0.01g/m1 NBT+0.005g/ml BCIP.
Sample preparation liquid is 1-5M, the carbonate buffer solution of pH9.6.
Lavation buffer solution is 0-0.05M, the phosphate buffer of pH7.4.
The detection method of described sword-leaved cymbidium leaf virus spot enzyme-linked assay kit is dot enzyme-linked detection method, and it may further comprise the steps:
1), reagent prepares: according to the needs that detect, dilute sample treating fluid and lavation buffer solution, standby;
2), sample preparation: get testing sample adding sample preparation liquid and fully grind centrifuging and taking supernatant, diluted for use again;
3), point sample: get the sample point sample on pvdf membrane after the processing, set up positive control and negative control simultaneously;
4), absorption: pvdf membrane is placed the dry 0-60min of 37 ℃ of constant temperature ovens, take out, with cleansing solution washing three times, 3min at every turn;
5), hatch I: get pvdf membrane and be soaked among the insulation liquid I, in 37 ℃ of insulation 30-90min, take out, with cleansing solution washing three times, 3min at every turn;
6), hatch II: get pvdf membrane and be soaked among the insulation liquid II, in 37 ℃ of insulation 30-90min.Take out, with cleansing solution washing three times, each 3min;
7), colour developing: the pvdf membrane after will washing takes out and is soaked in the colour developing liquid colour developing 510min;
8), detect and judge:, judge testing result according to detecting criterion.
After adopting such scheme, the present invention is by box body, is located at various in the box body and is assembled into economical and practical detection kit with liquid, pvdf membrane.The preparation method of kit is easy, quick, safe, reliable and economical and practical.Amount of samples is few, just can finish detection in the short period of time, and the result is easy to preserve, use this detection kit carry out method that dot enzyme-linked joint inspection surveys compare with other serologic test methods have simple, fast, characteristics such as sensitivity, high specificity, and have higher repeatability, a more advantage such as economical and practical, and kit uses and not to need specific apparatus, direct result of determination with the naked eye, and institute is so that apply in grass-roots unit.
Embodiment
The main agents that the present invention is used: the cymbidium mosaic virus polyclonal antibody is Xiamen Overseas Chinese Subtropical Plants Introduction Garden's preparation; PEG6000 (Macrogol 6000), 2 mercapto ethanol, Triton-100 (triton x-100), bovine serum albumin(BSA) (BSA) are available from Sigma company; Goat-anti rabbit enzyme labelled antibody, BCIP, NBT, DEAE ion-exchange filling material, IgG affinity chromatography filling material are available from Pierce company; Employed other conventional medicines and reagent are homemade analytical reagent in the test.
One, kit
Sword-leaved cymbidium leaf virus spot enzyme-linked assay kit mainly by box body, is located at various bottled in the box body and forms with liquid and pvdf membrane (PVDF membrane), various bottledly comprise with liquid:
1 bottle of insulation liquid I
1 bottle of insulation liquid II
1 bottle of colour developing damping fluid
1 of developer
1 of positive control
1 of negative control
1 bottle of sample preparation liquid
1 bottle of lavation buffer solution
Positive is the cymbidium mosaic virus virus of purifying, and its concentration is 100 μ g/ml;
Negative sample grinds the solution of healthy leaf preparation for sealing buffer solution.
Insulation liquid I is the phosphate buffer that contains cymbidium mosaic virus polyclonal antibody 0.05M, includes 1% gelatin.
Insulation liquid II is the phosphate buffer that contains goat-anti rabbit enzyme labelled antibody 0.05M, includes 1% gelatin.
The colour developing damping fluid is 0.1M, and pH9.5 contains the Tris damping fluid of MgCl2 10g/L+NaCl 2g/L.
Developer is 0.01g/ml NBT+0.005g/ml BCIP.
Sample preparation liquid is 2M, the carbonate buffer solution of pH9.6.
Lavation buffer solution is 0.01M, the phosphate buffer of pH7.4.
Two, preparation method
The preparation method of cymbidium mosaic virus detection kit of the present invention, it comprises the following steps:
1, the extraction of cymbidium mosaic virus and purifying: after getting the homogenate of the sick leaf adding of 100g datura 100ml0.5M citrate buffer solution (pH6.5), add the 100ml chloroform and stirred 30 minutes.At Beckman 10, centrifugal 20 minutes of 000rpm (JA-10).Get the NaCl that supernatant adds 4%PEG6000 and 0.3mol, left standstill after the stirring 1 hour.At Beckman 10, centrifugal 20 minutes of 000rpm (JA-10) abandons supernatant, fully repeats the PEG precipitation one time behind the dissolution precipitation with 0.5M citrate buffer solution (pH6.5).After getting for the second time the suspension of post precipitation usefulness 5mM borate buffer, at Beckman 10, centrifugal 20 minutes of 000rpm (JA-10) gets supernatant and uses P42A rotary head 40, centrifugal 2 hours of 000rpm in ultracentrifuge.Get precipitation and suspend again, carry out differential centrifugation again one time, get precipitation at last and dissolve with an amount of 0.01MPB (pH7.0) with 0.01MPB (pH7.0).If impurity is too many, carry out differential centrifugation again one time.Get thick purified virus and place 10%-40% sucrose pad gradient, use P42A rotary head 40, centrifugal 2 hours of 000rpm in ultracentrifuge.Collect viral band, use P42A rotary head 40 in ultracentrifuge once more, centrifugal 2 hours precipitation virus of 000rpm is dissolved among the 0.01MPB (pH7.0) at last, places-40 ℃ of preservations standby.
2, sero-fast preparation and purifying: select for use male rabbit to do immune animal, the purified virus that adds the emulsification of equivalent FreundShi Freund is done immunogene, adopts three intramuscular injection and twice intravenous immune rabbit.The amount of per injection virus is respectively 0.5,0.75,1,1.5 and 2mg, and be 7 days interval time, and last injection back blood sampling in 7-10 days 2 times is separated out serum and measured with agar double immunodiffusion method and tire.By the sad precipitation method in conjunction with DEAE ion-exchange chromatography antibody purification.Regulate antiserums to pH4.8 with 2 times of volume 0.1M ammonium acetates, press 0.75ml sad/1ml serum adds caprylic acid, mixes and stirs 1 hour, the centrifugal 30min of 5000rpm gets supernatant, places 10mM Tris-Cl pH8.5 damping fluid to spend the night bag filter, the desalination of dialysing.Get and dialyse completely that solution is splined in the ion exchange column, with 10mM Tris-HClpH8.5 damping fluid 5ml/min flushing 15min, carry out wash-out with the 10mMTris-HCl pH8.5 buffer concentration gradient that contains 0-1M NaCl again, collect the eluting peak part, concentrate dialysis, obtain the cymbidium mosaic virus polyclonal antibody of purifying.
3, the preparation of the positive, negative sample:
Positive is the cymbidium mosaic virus virus of purifying, and concentration is at 100 μ g/ml; Negative sample grinds the solution of healthy leaf preparation for sealing buffer solution.
4, various preparations with liquid:
Insulation liquid I is the phosphate buffer that contains cymbidium mosaic virus polyclonal antibody 0.05M, includes 1% gelatin;
Insulation liquid II is the phosphate buffer that contains goat-anti rabbit enzyme labelled antibody 0.05M, includes 1% gelatin;
The colour developing damping fluid is 0.1M, and pH9.5 contains the Tris of MgCl2 10g/L+NaCl 2g/L
Damping fluid;
Developer: be 0.01g/ml NBT+0.005g/ml BCIP;
Sample preparation liquid is 2M carbonate buffer solution (pH9.6);
Lavation buffer solution is 0.01M, the phosphate buffer of pH7.4;
Sealing buffer solution is 20-100mmol/L pH7.4 PBS+1-5% skimmed milk power.
Three, detection method embodiment
Dot enzyme-linked (DIBA) detection method of kit
1, reagent is prepared: according to the needs that detect, and dilute sample treating fluid and lavation buffer solution, standby;
2, sample preparation: get testing sample 0.1g adding 1ml sample preparation liquid and fully grind centrifuging and taking supernatant, diluted for use again;
3, point sample: get the sample point sample on pvdf membrane after 1 μ L handles, set up positive control and negative control simultaneously;
4, absorption: pvdf membrane is placed the dry 30min of 37 ℃ of constant temperature ovens, take out, with cleansing solution washing three times, each 3min;
5, hatch I: get pvdf membrane and be soaked among the insulation liquid I,, take out in 37 ℃ of insulation 45min, with cleansing solution washing three times, each 3min;
6, hatch II: get pvdf membrane and be soaked among the insulation liquid II, in 37 ℃ of insulation 45min.Take out, with cleansing solution washing three times, each 3min;
7, colour developing: the pvdf membrane after will washing takes out and is soaked in the colour developing liquid colour developing 5-10min;
8, detect judgement:, judge testing result according to detecting criterion.
Dot enzyme-linked detection criterion:
Under the situation that positive control colour developing, negative control do not develop the color, sample detection has the positive that is judged as of spot, otherwise negative.
Testing sample qualitatively judges the positive and negative of testing sample testing result by above standard.
For checking dot enzyme-linked detection kit to detect effect, detected 40 parts of orchid samples with this method, testing result is as follows:
Subordinate list 1 testing result
| RT-PCR(+) | RT-PCR(-) | Add up to | |
| DIBA (+) DIBA (-) adds up to | 30 1 31 | 1 8 9 | 31 9 40 |
Annotate: "+" represents positive; "-" represents negative
31 parts of samples by the RT-PCR detection positive, DIBA detects 30 parts of positive, 1 part of feminine gender;
9 parts of samples by RT-PCR detection feminine gender, DIBA detects negative sample part, 1 part of positive;
By testing result as can be known: the susceptibility of DIBA is 96.77%; Specificity 88.89%; Detection accuracy is 95%.
Claims (6)
1, sword-leaved cymbidium leaf virus spot enzyme-linked assay kit, mainly by box body, various bottled liquid and the pvdf membrane used that is located in the box body formed, and it is characterized in that:
Described various bottledly comprise with liquid:
1 bottle of insulation liquid I;
1 bottle of insulation liquid II;
1 bottle of colour developing damping fluid;
1 of developer;
1 of positive;
1 of negative sample;
1 bottle of sample preparation liquid;
1 bottle of lavation buffer solution.
2, sword-leaved cymbidium leaf virus spot enzyme-linked assay kit as claimed in claim 1 is characterized in that:
Described colour developing damping fluid is 0.05-0.3M, and pH9.5 contains MgCl
2The Tris damping fluid of 10g/L+NaCl2g/L;
Described developer is 0.01g/ml NBT+0.005g/ml BCIP;
Described sample preparation liquid is 1-5M, the carbonate buffer solution of pH9.6;
Described lavation buffer solution is 0-0.05M, the phosphate buffer of pH7.4.
3, sword-leaved cymbidium leaf virus spot enzyme-linked assay kit as claimed in claim 1 or 2 is characterized in that:
Described insulation liquid I is the 0.01-0.1M phosphate buffer that contains the cymbidium mosaic virus polyclonal antibody, includes 1% gelatin;
Described insulation liquid II is the 0.01-0.1M phosphate buffer that contains goat-anti rabbit enzyme labelled antibody, includes 1% gelatin;
Described positive is the cymbidium mosaic virus virus of purifying;
Described negative sample grinds the solution of healthy leaf preparation for sealing buffer solution.
4, as claims 3 described sword-leaved cymbidium leaf virus spot enzyme-linked assay kits, it is characterized in that: described positive is the cymbidium mosaic virus of purifying, and concentration is 0.1 μ g-100 μ g/ml.
5, the preparation method of sword-leaved cymbidium leaf virus spot enzyme-linked assay kit as claimed in claim 1 is characterized in that: described preparation method mainly comprises the following steps:
1), the extraction and the purifying of cymbidium mosaic virus: after getting the sick leaf of 100g datura and adding 100ml0.5M citrate buffer solution (pH6.5) homogenate, add the 100ml chloroform and stirred 30 minutes, at Beckman 10, centrifugal 20 minutes of 000rpm (JA-10); Get the NaCl that supernatant adds 4%PEG6000 and 0.3mol, left standstill after the stirring 1 hour, at Beckman 10, centrifugal 20 minutes of 000rpm (JA-10), abandon supernatant, after repeating one time the PEG precipitation behind the abundant dissolution precipitation of 0.5M citrate buffer solution (pH6.5), get post precipitation for the second time to use the suspension of 5mM borate buffer, at Beckman 10, centrifugal 20 minutes of 000rpm (JA-10), get supernatant and use P42A rotary head 40 in ultracentrifuge, centrifugal 2 hours of 000rpm gets precipitation and suspends again with 0.01M PB (pH7.0), carry out differential centrifugation again one time, get precipitation at last with an amount of 0.01M PB (pH7.0) dissolving,, carry out differential centrifugation again one time if impurity is too many, get thick purified virus and place 10%-40% sucrose pad gradient, use P42A rotary head 40 in ultracentrifuge, centrifugal 2 hours of 000rpm collects viral band, use P42A rotary head 40 in ultracentrifuge once more, centrifugal 2 hours precipitation virus of 000rpm is dissolved among the 0.01M PB (pH7.0) at last, places-40 ℃ of preservations standby;
2), sero-fast preparation and purifying: select for use male rabbit to do immune animal, the purified virus that adds the emulsification of equivalent FreundShi Freund is done immunogene, adopt three intramuscular injection and twice intravenous immune rabbit, the amount of per injection virus is respectively 0.5,0.75,1,1.5 and 2mg, be 7 days interval time, last injection back blood sampling in 7-10 days 2 times, separating out serum tires with agar double immunodiffusion method mensuration, by the sad precipitation method in conjunction with DEAE ion-exchange chromatography antibody purification, regulate antiserum to pH4.8 with 2 times of volume 0.1M ammonium acetates, press 0.75ml sad/1ml serum adds caprylic acid, mix and stirred 1 hour, the centrifugal 30min of 5000rpm, get supernatant, place 10mM Tris-Cl pH8.5 damping fluid to spend the night bag filter, the dialysis desalination, get and dialyse completely that solution is splined in the ion exchange column, with 10mM Tris-HClpH8.5 damping fluid 5ml/min flushing 15min, carry out wash-out with the 10mMTris-HCl pH8.5 buffer concentration gradient that contains 0-1M NaCl again, collect the eluting peak part, concentrate dialysis, obtain the cymbidium mosaic virus polyclonal antibody of purifying;
3), the preparation of the positive, negative sample:
Positive is the cymbidium mosaic virus virus of purifying, and concentration is in 0.1 μ g-100 μ g/ml scope; Negative sample grinds the solution of healthy leaf preparation for sealing buffer solution;
4), various preparations with liquid:
Insulation liquid I is the 0.01-0.1M phosphate buffer that contains the cymbidium mosaic virus polyclonal antibody, includes 1% gelatin;
Insulation liquid II is the 0.01-0.1M phosphate buffer that contains goat-anti rabbit enzyme labelled antibody, includes 1% gelatin;
The colour developing damping fluid is 0.05-0.3M, and pH9.5 contains MgCl
2The Tris damping fluid of 10g/L+NaCl 2g/L;
Developer is 0.01g/ml NBT+0.005g/ml BCIP;
Sample preparation liquid is 1-5M, the carbonate buffer solution of pH9.6;
Lavation buffer solution is 0-0.05M, the phosphate buffer of pH7.4.
6, the detection method of sword-leaved cymbidium leaf virus spot enzyme-linked assay kit as claimed in claim 1 is characterized in that: described detection method is dot enzyme-linked detection method, and it may further comprise the steps:
1), reagent prepares: according to the needs that detect, dilute sample treating fluid and lavation buffer solution, standby;
2), sample preparation: get testing sample adding sample preparation liquid and fully grind centrifuging and taking supernatant, diluted for use again;
3), point sample: get the sample point sample on pvdf membrane after the processing, set up positive control and negative control simultaneously;
4), absorption: pvdf membrane is placed the dry 0-60min of 37 ℃ of constant temperature ovens, take out, with cleansing solution washing three times, 3min at every turn;
5), hatch I: get pvdf membrane and be soaked among the insulation liquid I, in 37 ℃ of insulation 30-90min, take out, with cleansing solution washing three times, 3min at every turn;
6), hatch II: get pvdf membrane and be soaked among the insulation liquid II, in 37 ℃ of insulation 30-90min, take out, with cleansing solution washing three times, 3min at every turn;
7), colour developing: the pvdf membrane after will washing takes out and is soaked in the colour developing liquid colour developing 5-10min;
8), detect and judge:, judge testing result according to detecting criterion.
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| CN 200610164434 CN1975428A (en) | 2006-02-10 | 2006-12-08 | Sword-leaved cymbidium leaf virus spot enzyme-linked assay kit, preparation thereof and detecting method |
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| CN200610033609 | 2006-02-10 | ||
| CN200610033609.4 | 2006-02-10 | ||
| CN 200610164434 CN1975428A (en) | 2006-02-10 | 2006-12-08 | Sword-leaved cymbidium leaf virus spot enzyme-linked assay kit, preparation thereof and detecting method |
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102043056A (en) * | 2010-11-10 | 2011-05-04 | 福建省亚热带植物研究所 | Preparation method for immune colloidal gold test strip for simultaneously detecting orchid viruses CyMV and ORSV |
| CN102520185A (en) * | 2011-11-18 | 2012-06-27 | 河南省农业科学院 | Serologic detecting kit for potyvirus on sweet potato and detecting method thereof |
| CN103760346A (en) * | 2014-01-25 | 2014-04-30 | 云南省农业科学院生物技术与种质资源研究所 | Dot fluorescence immunoassay method for quantitatively detecting plant virus |
| CN108220253A (en) * | 2017-10-09 | 2018-06-29 | 南京林业大学 | It is a kind of to detach method viral in Bursaphelenchus xylophilus body |
| CN108593633A (en) * | 2018-04-19 | 2018-09-28 | 中山大学 | A kind of Test paper for quickly detecting saliva uric acid |
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2006
- 2006-12-08 CN CN 200610164434 patent/CN1975428A/en active Pending
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102043056A (en) * | 2010-11-10 | 2011-05-04 | 福建省亚热带植物研究所 | Preparation method for immune colloidal gold test strip for simultaneously detecting orchid viruses CyMV and ORSV |
| CN102520185A (en) * | 2011-11-18 | 2012-06-27 | 河南省农业科学院 | Serologic detecting kit for potyvirus on sweet potato and detecting method thereof |
| CN103760346A (en) * | 2014-01-25 | 2014-04-30 | 云南省农业科学院生物技术与种质资源研究所 | Dot fluorescence immunoassay method for quantitatively detecting plant virus |
| CN103760346B (en) * | 2014-01-25 | 2015-06-24 | 云南省农业科学院生物技术与种质资源研究所 | Dot fluorescence immunoassay method for quantitatively detecting plant virus |
| CN108220253A (en) * | 2017-10-09 | 2018-06-29 | 南京林业大学 | It is a kind of to detach method viral in Bursaphelenchus xylophilus body |
| CN108220253B (en) * | 2017-10-09 | 2019-11-05 | 南京林业大学 | A method of it is viral in separation Bursaphelenchus xylophilus body |
| CN108593633A (en) * | 2018-04-19 | 2018-09-28 | 中山大学 | A kind of Test paper for quickly detecting saliva uric acid |
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