CN102095855A - Insecticidal crystal protein CrylAc enzyme linked immunosorbent detection kit - Google Patents
Insecticidal crystal protein CrylAc enzyme linked immunosorbent detection kit Download PDFInfo
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- CN102095855A CN102095855A CN201010601129XA CN201010601129A CN102095855A CN 102095855 A CN102095855 A CN 102095855A CN 201010601129X A CN201010601129X A CN 201010601129XA CN 201010601129 A CN201010601129 A CN 201010601129A CN 102095855 A CN102095855 A CN 102095855A
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- cry1ac
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Abstract
The invention discloses an insecticidal crystal protein CrylAc enzyme linked immunosorbent detection kit. In the CrylAc protein detection of the kit, the lowest detection limit is 7.81ng/mL, the linear detection range is 62.5-500ng/mL, the linear interval fitted equation y is equal to 1.1136Ln(x)-4.164, and R2 is equal to 0.9991. No cross reaction occurs among the kit and CrylAb and Crylc proteins. The kit is suitable for qualitative detection of the CrylAc protein in transgenic plant leaves, fruits and derivatives. The kit can detect large batch of samples simultaneously, is convenient and fast, has very important realistic significance in analyzing related transgenic articles, simultaneously ensures the detection cost to be greatly reduced and has potential economic value.
Description
Technical field
The present invention relates to kit, relate in particular to a kind of insecticidal crystal protein Cry1Ac enzyme-linked immuno sorbent assay kit.
Technical background:
In the genetically modified crops, especially with maximum to the research of Bt-Cry transgene protein.The Bt gene is a kind of anti insect gene that research at present is most widely used, and the effect aspect pest control is more and more important, and the Bt toxalbumin has higher resistance to 8 kinds of paddy rice lepidoptera pests such as striped rice borer, yellow rice borer, pink rice borer, rice leaf roller, rice leaf feeders.It is reported that known Bt anti insect gene has reached 70 kinds at present, research of Bt anti-pest crop and application are just at high speed development.Carrying out enetically modified food research and development and business-like while, for the security to enetically modified food gives comprehensive assessment, make the consumer can distinguish enetically modified food and wholefood fast, develop corresponding transgene component quick detection kit and be of great practical significance.Cry1Ac albumen is a kind of of Bt toxalbumin, and the Cry1Ac protein detection kit meaning of therefore developing a kind of simple rapid sensitive is great equally.
Common Bt method of protein detection has based on the PCR method of nucleic acid DNA detection with based on the Protein Detection enzyme linked immunosorbent assay at present.The PCR method can begin to the ripe whole process of crop transgene component accurately to be detected from crop seed, be not subjected to the influence of the disposal route of sample, but problems such as false positive and false negative appear easily, for transgenic product through highly handling and derivant thereof, as vegetable oil, sugar or starch etc., they do not contain any DNA composition, so PCR method is difficult to realize to this type of methods for detecting transgenic foods; Enzyme linked immunological absorption detects and is based on that antigen combine with the specificity of antibody and a kind of Fast Detection Technique of the efficient catalytic characteristic of zymolyte, has fast, characteristics such as sensitivity, accurate, low cost, can be qualitative again can detection by quantitative target transgene protein.Compare the detection of nucleic acid level, the detection of protein level is science, more accurate more.
The conventional mouse monoclonal antibody of rabbit monoclonal antibodies has more advantages.At first, rabbit can discern than mouse and more many epitope antigen determinant of immunizing antigen.Secondly,, can carry out more Fusion of Cells experiment, make the positive fused cell of high flux screening become possibility because the rabbit spleen is bigger.
Summary of the invention
The objective of the invention is to overcome the deficiencies in the prior art, a kind of have high specific, highly sensitive insecticidal crystal protein Cry1Ac enzyme-linked immuno sorbent assay kit are provided.
Insecticidal crystal protein Cry1Ac enzyme-linked immuno sorbent assay kit comprises box body and is located at 96 interior hole ELISA Plate and reagent of box body.96 hole ELISA Plate endoperidiums have anti-Cry1Ac monoclonal antibody, and the reagent in the box is as follows:
(1) concentrated solution for washing;
(2) dilution concentrate;
(3) Cry1Ac protein standard solution;
(4) horseradish peroxidase is marked anti-Cry1Ac polyclonal antibody;
(5) substrate dilution;
(6) substrate;
(7) substrate colour developing liquid;
(8) reaction terminating liquid.
Described anti-Cry1Ac rabbit monoclonal antibodies and rabbit polyclonal antibody all can with Cry1Ac albumen specific bond; Described concentrated solution for washing contains sodium chloride 7～9 g, potassium dihydrogen phosphate 0.1～0.3 g, sodium hydrogen phosphate 3～5 g, potassium chloride 0.1～0.3 g, polysorbas20 0.5～1mL and deionized water; Described dilution concentrate is for containing sodium chloride 7～9 g, potassium dihydrogen phosphate 0.1～0.3 g, sodium hydrogen phosphate 3～5 g, potassium chloride 0.1～0.3 g and deionized water; Described anti-Cry1Ac albumen rabbit monoclonal antibodies is to be immune animal with the rabbit, and the process hybridoma technology finally obtains, and anti-Cry1Ac albumen rabbit polyclonal antibody is to be immune animal with the rabbit, and animal blood serum obtains through protein A gel column purifying; Described horseradish peroxidase enzyme mark polyclonal antibody is the product of horseradish peroxidase and the coupling of anti-Cry1Ac rabbit polyclonal antibody; Described substrate colour developing liquid is 3,3,5,5-tetramethyl benzidine preparation liquid; Described substrate dilution is the pH5.0 citrate buffer solution, contains 3～6 g citric acids, 6～9g sodium hydrogen phosphate and deionized water in the prescription; Described Cry1Ac protein standard solution concentration is respectively 2000ng/mL, 1000ng/mL, 500ng/mL, 250ng/mL, 125ng/mL, 62.5 ng/mL, 31.25 ng/mL, 15.62 ng/mL, 7.81 ng/mL, 3.90 ng/mL; Described substrate is hydrogen peroxide or urea peroxide; Described reaction terminating liquid is a 2M sulfuric acid.
The present invention is in the testing process of Cry1Ac albumen, and its lowest detection is limited to 7.81ng/mL, and its linear detection range is 62.5 ~ 500ng/mL, fit equation y=1.1136Ln (x)-4.164 between linear zone, R
2=0.9991.
The present invention is suitable for the detection of Cry1Ac albumen in rotaring gene plant blade, fruit and the derivant thereof.This kit can detect sample in enormous quantities simultaneously, and is convenient quick again, and the analysis of transgenosis relative article is of great practical significance; The detection cost is reduced greatly, have potential economic worth.
Description of drawings
Fig. 1 is a Cry1Ac albumen double antibodies sandwich enzyme linked immunological adsorption criteria curve;
Fig. 2 is kit and Cry1Ab, Cry1c albumen cross reaction result.
Concrete embodiment
The present invention is based on that antibody combines with antigentic specificity and the insecticidal crystal protein Cry1Ac euzymelinked immunosorbent assay (ELISA) developed absorption detection kit, and this kit can detect Cry1Ac albumen in rotaring gene plant blade, fruit and the derivant thereof based on immune response and enzymatic reaction.Insecticidal crystal protein Cry1Ac enzyme-linked immuno sorbent assay kit comprises box body and is located at box body interior 96 hole ELISA Plate and reagent 96 hole ELISA Plate endoperidiums anti-Cry1Ac monoclonal antibody, and the reagent in the box comprises (1) concentrated solution for washing; (2) dilution concentrate; (3) Cry1Ac protein standard solution; (4) horseradish peroxidase is marked anti-Cry1Ac polyclonal antibody; (5) substrate dilution; (6) substrate; (7) substrate colour developing liquid; (8) reaction terminating liquid.
Described coated antibody can with Cry1Ac albumen specific bond, horseradish peroxidase-labeled how anti-equally can with Cry1Ac albumen specific bond.Described concentrated solution for washing contains sodium chloride 7～9 g, potassium dihydrogen phosphate 0.1～0.3 g, sodium hydrogen phosphate 3～5 g, potassium chloride 0.1～0.3 g, polysorbas20 0.5～1mL and deionized water; Described dilution concentrate is for containing sodium chloride 7～9 g, potassium dihydrogen phosphate 0.1～0.3 g, sodium hydrogen phosphate 3～5 g, potassium chloride 0.1～0.3 g and deionized water; Described anti-Cry1Ac albumen rabbit monoclonal antibodies is to be immune animal with the rabbit, and the process hybridoma technology finally obtains, and anti-Cry1Ac albumen rabbit polyclonal antibody is to be immune animal with the rabbit, and animal blood serum obtains through protein A gel column purifying; Described horseradish peroxidase (HRP) enzyme mark polyclonal antibody is the product of horseradish peroxidase (HRP) and the coupling of anti-Cry1Ac rabbit polyclonal antibody; Described substrate colour developing liquid is 3,3,5,5-tetramethyl benzidine preparation liquid; Described substrate dilution is the pH5.0 citrate buffer solution, contains 3～6 g citric acids, 6～9g sodium hydrogen phosphate and deionized water in the prescription; Described Cry1Ac protein standard solution concentration is respectively 2000ng/mL, 1000ng/mL, 500ng/mL, 250ng/mL, 125ng/mL, 62.5 ng/mL, 31.25 ng/mL, 15.62 ng/mL, 7.81 ng/mL, 3.90 ng/mL; Described substrate is hydrogen peroxide or urea peroxide; Described reaction terminating liquid is a 2M sulfuric acid.
Reagent setting in the box:
(1) concentrated solution for washing is 1 bottle, and 25～35mL/ bottle contains sodium chloride 7～9g, potassium dihydrogen phosphate 0.1～0.3 g, sodium hydrogen phosphate 3～5 g, potassium chloride 0.1～0.3 g, polysorbas20 0.5～1mL, is 30～40 times of normal working concentration; (2) 1 bottle of concentrate of dilution, 25～35mL/ bottle contains sodium chloride 7～9g, potassium dihydrogen phosphate 0.1～0.3 g, sodium hydrogen phosphate 3～5 g, potassium chloride 0.1～0.3 g, is 30～40 times of normal working concentration; (3) Cry1Ac protein standard solution (2000ng/mL, 1000ng/mL, 500ng/mL, 250ng/mL, 125ng/mL, 62.5 ng/mL, 31.25 ng/mL, 15.62 ng/mL, 7.81 ng/mL, 3.90 ng/mL) is each one bottle, the 1mL/ bottle; (4) horseradish peroxidase mark Cry1Ac polyclonal antibody one pipe, 400 μ g/ pipe, dilution is 160 times during use; (5) the substrate dilution is 1 bottle, and 25～35mL/ bottle includes 3～6g citric acid, 6～9g sodium hydrogen phosphate, for the 30-40 of normal use amount doubly; (6) substrate colour developing liquid is 1 bottle, the 1mL/ bottle; (7) substrate colour developing liquid is 1 bottle of tetramethyl benzidine preparation liquid, the 4mL/ bottle, and compound method is that 10mgTMB is dissolved among the 2mLDMSO; (8) reaction terminating liquid is 1 bottle, 15～20mL/ bottle, and it is a 2M sulfuric acid.
Embodiment 1: the generation of insecticidal crystal protein Cry1Ac rabbit monoclonal antibodies
1.Cry1Ac protein immunization rabbit
The Cry1Ac albumen of 500 μ g is mixed with the equivalent Freund's complete adjuvant, fully after the emulsification as immunogene, at 5 injecting immunes of the rabbit subcutaneous branch in (three new zealand white rabbits) back, in beginning the 14th, 28,42,56 day after the immunity, the Cry1Ac albumen with 200 μ g then carries out immunity with identical method with the incomplete Freund mixing respectively.
2. Fusion of Cells
Rabbit after the 5th immunity is got blood and isolates serum and carry out immunizing potency and measure, and the higher rabbit of selecting to tire enters Fusion of Cells.Get the rabbit spleen and carried out Fusion of Cells preceding ten days, rabbit is carried out booster immunization one time, get aseptic spleen then.Spleen cell and myeloma cell that selection is in exponential phase are merged, and fusion agent is 50% PEG, and HAT is as selective medium, and are laid in 40 96 orifice plates and cultivate.
3. hybridoma screening
Carried out primary dcreening operation to cloning son on the 17th day after the Fusion of Cells, filter out positive colony by indirect enzyme-linked immunosorbent absorption, and the cell line of correspondence transferred in the 24 porocyte culture plates cultivated 7 days, filter out the secretion high-affinity antibody by indirect enzyme-linked immunosorbent absorption then, and clone's that cell growth state is good, select three clones to carry out the plasmid reorganization, and final production go out monoclonal antibody.
Embodiment 2: the generation of insecticidal crystal protein Cry1Ac rabbit polyclonal antibody
The Cry1Ac albumen of 500 μ g is mixed with the equivalent Freund's complete adjuvant, fully after the emulsification as immunogene, at 5 injecting immunes of the rabbit subcutaneous branch in (two new zealand white rabbits) back, in beginning the 14th, 28,42,56 day after the immunity, the Cry1Ac albumen with 200 μ g then carries out immunity with identical method with the incomplete Freund mixing respectively.Rabbit after the 5th immunity is got blood and isolates serum and carry out immunizing potency and measure, and the higher rabbit of selecting to tire carries out booster immunization twice, obtains the rabbit whole blood then.The rabbit whole blood is isolated serum earlier, obtains anti-Cry1Ac rabbit polyclonal antibody through protein A gel column purifying then.
Embodiment 3: the horseradish peroxidase-labeled of anti-Cry1Ac rabbit polyclonal antibody
A. the processing of antibody: 2mg antibody is dissolved in CBS (50mM, pH 9.6), and 4 ℃, dialysed overnight in the CBS solution, liquid is changed 2 times in the centre.
B. the oxidation of horseradish peroxidase (following steps are all carried out under the lucifuge condition):
2mg HRP+0.4mL ddH
2The O mixing;
10mg NaIO
4+ 5mL ddH
2The O mixing;
Get NaIO
4(aq) 45 μ L slowly add among the HRP (aq), and the vibration mixing is in room temperature lucifuge reaction 20min;
Get ethylene glycol 40 μ L and join in the above-mentioned solution, mixing, room temperature continues lucifuge reaction 30min;
C. crosslinked: as the HRP of oxidation to be joined in the antibody bag filter mixing, 4 ℃ of lucifuge dialysis (50mM CBS) 2h;
D. reduction: take out product after crosslinked as in the light resistant container, add 45 μ L NaBH then
4Solution (5mg NaBH
4+ 0.5mL ddH
2O), mixing, 4 ℃ of lucifuge reaction 2h, the centre shakes up 2 ~ 3 times;
E. dialysis: above-mentioned solution is placed PBS(10mM, pH7.2) more than the dialysis 18h, 4 ℃, change liquid 3 ~ 4 times
F. gather in the crops the good antibody of mark-HRP solution, standby (equal-volume glycerine is stored in-20 ℃)
The monoclonal antibody bag quilt of embodiment 4:96 hole ELISA Plate
Anti-Cry1Ac rabbit monoclonal antibodies is 5 μ g/mL with the CBS dilution, 100 μ L/ holes join in the ELISA Plate hole, and leave standstill in 37 ℃ and to hatch 2h, the coated antibody that inclines, PBST cleansing solution washing 3 times pats dry, in every hole, add 200 μ L3% skimmed milk powers then, 37 ℃ were sealed 1 hour, took out back washing 3～5 times, and drying is preserved after sealing film.
Embodiment 5: the kit principle of work
The monoclonal antibody that is coated on the ELISA Plate can combine with a certain specific site on the target protein specifically, washes the unnecessary albumen that not can be incorporated on the coated antibody off, and enzyme labelled antibody can combine with the albumen on being attached to coated antibody.The binding capacity of enzyme labelled antibody becomes positive correlation with the amount of the interior albumen of plate, and through process color, the amount that the enzyme labelled antibody in the plate is stayed in the dark more explanation of color is many more, and promptly the content of target protein is high more.Make typical curve with protein standard solution in the mensuration process, the content of target protein in the working sample simultaneously is by colour developing OD value and the typical curve content to target protein in the judgement sample recently.
Embodiment 6: the pre-treatment of testing sample
A. plant leaf blade etc. contains the processing of high fibre solid material
Blade etc. is dried in cleaning put into mortar, pour liquid nitrogen into and rapidly it is ground to Powdered, transfer in the centrifuge tube, add PBST or Na
2CO
3Protein extracts such as damping fluid are with its abundant mixing, and 30min is hatched in the room temperature vibration.Then in 4 ℃, the centrifugal 10min of 5000rpm.Supernatant is protein extract;
B. the processing of fluid sample
With fluid sample be directly used in measure or the dilution suitable multiple after measure.
The use of embodiment 7:Cry1Ac protein detection kit
(1) reagent preparation
A. dilute concentrate: the dilution concentrate uses with behind 30～40 times of ultrapure water or the distilled water dilutings in the kit;
B. concentrated solution for washing: concentrated solution for washing uses with behind 30～40 times of ultrapure water or the distilled water dilutings in the kit;
C. substrate dilution: the substrate dilution uses with behind 30～40 times of ultrapure water or the distilled water dilutings in the kit.
(2) the kit operating process is as follows:
A. kit needs ambient-temp-stable more than half an hour before using;
B. take out ELISA Plate, add each 100 μ L of standard specimen and sample in corresponding hole, standard specimen and sample all can be done 2 ~ 3 repetitions, and hatch 1h in 37 ℃ of vibrations;
C. pour out the liquid in the hole, microwell plate is inverted on the thieving paper pats,,, pat dry with 250 μ L cleansing solutions washing 3～5 times to guarantee to remove fully the liquid in the hole;
D. add with PBST and dilute good enzyme labelled antibody 100 μ L/ holes, and hatch 1h in 37 ℃ of vibrations;
E. pour out the liquid in the hole, microwell plate is inverted on the thieving paper pats,,, pat dry with 250 μ L cleansing solutions washing 5 times to guarantee to remove fully the liquid in the hole;
F. get substrate colour developing liquid 400 μ L and join in the 10mL substrate dilution, and add 4 μ L substrate hydrogen peroxide, after the vibration evenly, every hole adds the chromophoric solution for preparing more than the 100 μ L, slight vibration plate, and 10min is hatched in 37 ℃ of dark places;
G. add 50 μ L stop buffers, measure OD
450The value result of determination.
Embodiment 8: the enzyme-linked immunosorbent assay kit result
1. enzyme linked immunological absorption examination criteria curve and sensitivity
Logarithm with Cry1Ac protein solution concentration is made horizontal ordinate, with OD
450Make ordinate.As lowest detectable limit, has the linear detection range that the good linear section is this kit with the protein concentration of the value correspondence of the mean value+3 times blank standard deviation of blank on the curve.From Fig. 1 .Cry1Ac albumen double antibodies sandwich enzyme linked immunological adsorption criteria curve as can be known the linear detection range of this kit be 61.25 ~ 500ng/mL, fit equation y=1.1136Ln (x)-4.164 between its linear zone, R
2=0.9991. is limited to 7.81ng/ml. by calculating this kit to the lowest detection of Cry1Ac albumen
2. the cross reaction between enzyme linked immunological absorption detection architecture and other Cry albumen
Respectively with the logarithm of Cry1Ac, Cry1Ab, Cry1c protein solution concentration as horizontal ordinate, with OD
450Make ordinate.From Fig. 2. this enzyme linked immunological absorption detection architecture is to Cry1Ab, Cry1c albumen no cross reaction as can be known.
Claims (10)
1. an insecticidal crystal protein Cry1Ac enzyme-linked immuno sorbent assay kit is characterized in that comprising box body and is located at 96 interior hole ELISA Plate and reagent of box body, and 96 hole ELISA Plate endoperidiums have anti-Cry1Ac monoclonal antibody, and the reagent in the box is as follows:
(1) concentrated solution for washing;
(2) dilution concentrate;
(3) Cry1Ac protein standard solution;
(4) horseradish peroxidase is marked anti-Cry1Ac polyclonal antibody;
(5) substrate dilution;
(6) substrate;
(7) substrate colour developing liquid;
(8) reaction terminating liquid.
2. a kind of insecticidal crystal protein Cry1Ac enzyme-linked immuno sorbent assay kit according to claim 1, it is characterized in that described anti-Cry1Ac albumen rabbit monoclonal antibodies and rabbit polyclonal antibody all can with Cry1Ac albumen specific bond, monoclonal antibody is 2.69 * 10 to the affinity costant of Cry1Ac albumen
9M
-1
3. a kind of insecticidal crystal protein Cry1Ac enzyme-linked immuno sorbent assay kit according to claim 1 is characterized in that described concentrated solution for washing contains sodium chloride 7～9 g, potassium dihydrogen phosphate 0.1～0.3 g, sodium hydrogen phosphate 3～5 g, potassium chloride 0.1～0.3 g, polysorbas20 0.5～1mL and deionized water.
4. a kind of insecticidal crystal protein Cry1Ac enzyme-linked immuno sorbent assay kit according to claim 1 is characterized in that described dilution concentrate is for containing sodium chloride 7～9 g, potassium dihydrogen phosphate 0.1～0.3 g, sodium hydrogen phosphate 3～5 g, potassium chloride 0.1～0.3 g and deionized water.
5. a kind of insecticidal crystal protein Cry1Ac enzyme-linked immuno sorbent assay kit according to claim 1, it is characterized in that described anti-Cry1Ac albumen rabbit monoclonal antibodies is to be immune animal with the rabbit, finally obtain through hybridoma technology, anti-Cry1Ac albumen rabbit polyclonal antibody is to be immune animal with the rabbit, and animal blood serum obtains through Sepharose CL-4B Protein A gel column purifying.
6. a kind of insecticidal crystal protein Cry1Ac enzyme-linked immuno sorbent assay kit according to claim 1 is characterized in that described horseradish peroxidase enzyme mark polyclonal antibody is the product of horseradish peroxidase and the coupling of anti-Cry1Ac rabbit polyclonal antibody.
7. insecticidal crystal protein Cry1Ac enzyme-linked immuno sorbent assay kit according to claim 1 is characterized in that described substrate colour developing liquid is 3,3,5,5-tetramethyl benzidine preparation liquid.
8. a kind of insecticidal crystal protein Cry1Ac enzyme-linked immuno sorbent assay kit according to claim 1, it is characterized in that described substrate dilution is the pH5.0 citrate buffer solution, contain 3～6 g citric acids, 6～9g sodium hydrogen phosphate and deionized water in the prescription.
9. a kind of insecticidal crystal protein Cry1Ac enzyme-linked immuno sorbent assay kit according to claim 1, it is characterized in that described Cry1Ac protein standard solution concentration is respectively 2000ng/mL, 1000ng/mL, 500ng/mL, 250ng/mL, 125ng/mL, 62.5 ng/mL, 31.25 ng/mL, 15.62 ng/mL, 7.81 ng/mL, 3.90 ng/mL.
10. a kind of insecticidal crystal protein Cry1Ac enzyme-linked immuno sorbent assay kit according to claim 1 is characterized in that described substrate is hydrogen peroxide or urea peroxide; Described reaction terminating liquid is a 2M sulfuric acid.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102495211A (en) * | 2011-12-07 | 2012-06-13 | 河北省科学院生物研究所 | Beta-fructofuranosidase enzyme-linked immunoassay detection kit |
| CN102590527A (en) * | 2012-01-16 | 2012-07-18 | 中国农业大学 | Method for detecting insecticidal crystal proteins Bt Cry1Ab/Ac and special enzyme linked immunosorbent assay kit thereof |
| CN103116027A (en) * | 2011-11-16 | 2013-05-22 | 北京市理化分析测试中心 | Double-antibody sandwich ELISA detection method of transgenic plant BtCry1Ac protein |
| CN103197075A (en) * | 2012-01-16 | 2013-07-10 | 华中农业大学 | Method for detecting Bt protein in transgenic rice by quantum dot |
| CN103278632A (en) * | 2013-04-26 | 2013-09-04 | 江苏省农业科学院 | ELISA kit for fast detection of corn phosphoenolpyruvate carboxylase (PEPC) gene transgenic crop and use thereof |
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| CN108291233A (en) * | 2015-09-09 | 2018-07-17 | 先正达参股股份有限公司 | composition and method for protein detection |
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| CN1501082A (en) * | 2002-11-18 | 2004-06-02 | 中国农业科学院原子能利用研究所 | Bt crystallin CrylAc radio-immunity test reagent box, preparing and detecting method thereof |
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| CN1501082A (en) * | 2002-11-18 | 2004-06-02 | 中国农业科学院原子能利用研究所 | Bt crystallin CrylAc radio-immunity test reagent box, preparing and detecting method thereof |
| CN101699292A (en) * | 2009-10-29 | 2010-04-28 | 浙江大学 | Rabbit monoclonal antibody based ciprofloxacin residue analysis enzyme-linked immune adsorption kit |
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| CN103116027A (en) * | 2011-11-16 | 2013-05-22 | 北京市理化分析测试中心 | Double-antibody sandwich ELISA detection method of transgenic plant BtCry1Ac protein |
| CN102495211A (en) * | 2011-12-07 | 2012-06-13 | 河北省科学院生物研究所 | Beta-fructofuranosidase enzyme-linked immunoassay detection kit |
| CN102495211B (en) * | 2011-12-07 | 2015-05-20 | 河北省科学院生物研究所 | Beta-fructofuranosidase enzyme-linked immunoassay detection kit |
| CN102590527A (en) * | 2012-01-16 | 2012-07-18 | 中国农业大学 | Method for detecting insecticidal crystal proteins Bt Cry1Ab/Ac and special enzyme linked immunosorbent assay kit thereof |
| CN103197075A (en) * | 2012-01-16 | 2013-07-10 | 华中农业大学 | Method for detecting Bt protein in transgenic rice by quantum dot |
| CN102590527B (en) * | 2012-01-16 | 2014-04-23 | 中国农业大学 | Method for detecting insecticidal crystal proteins Bt Cry1Ab/Ac and special enzyme linked immunosorbent assay kit thereof |
| CN103278632A (en) * | 2013-04-26 | 2013-09-04 | 江苏省农业科学院 | ELISA kit for fast detection of corn phosphoenolpyruvate carboxylase (PEPC) gene transgenic crop and use thereof |
| CN105137061B (en) * | 2015-07-28 | 2016-09-14 | 环境保护部南京环境科学研究所 | A kind of soil remains the original position ELISA quantitative determination method of bt albumen |
| CN108291233A (en) * | 2015-09-09 | 2018-07-17 | 先正达参股股份有限公司 | composition and method for protein detection |
| CN108291233B (en) * | 2015-09-09 | 2021-11-16 | 先正达参股股份有限公司 | Compositions and methods for protein detection |
| CN109781992A (en) * | 2018-12-27 | 2019-05-21 | 中国农业科学院生物技术研究所 | A kind of colloidal gold immunochromatographimethod quick measuring card of insect resistance protein Cry1C |
| CN109781992B (en) * | 2018-12-27 | 2021-04-06 | 中国农业科学院生物技术研究所 | Colloidal gold immunochromatographic assay rapid test card for insect-resistant protein Cry1C |
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Application publication date: 20110615 |