CN106596948A - Time-resolved fluoroimmunoassy kit for detecting dozine - Google Patents
Time-resolved fluoroimmunoassy kit for detecting dozine Download PDFInfo
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- CN106596948A CN106596948A CN201510664248.2A CN201510664248A CN106596948A CN 106596948 A CN106596948 A CN 106596948A CN 201510664248 A CN201510664248 A CN 201510664248A CN 106596948 A CN106596948 A CN 106596948A
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/577—Immunoassay; Biospecific binding assay; Materials therefor involving monoclonal antibodies binding reaction mechanisms characterised by the use of monoclonal antibodies; monoclonal antibodies per se are classified with their corresponding antigens
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/531—Production of immunochemical test materials
- G01N33/532—Production of labelled immunochemicals
- G01N33/533—Production of labelled immunochemicals with fluorescent label
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2430/00—Assays, e.g. immunoassays or enzyme assays, involving synthetic organic compounds as analytes
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Abstract
The invention discloses a time-resolved fluoroimmunoassy kit for detecting dozine. The time-resolved fluoroimmunoassy kit for detecting dozine is composed of a porous coated plate, a buffer solution, a dozine standard substance, a lyophilized antibody product for dozine, a sheep-anti-mouse antibody labeled with europium, a washing solution and an enhancement solution. The detection method of the time-resolved fluoroimmunoassy kit for detecting dozine includes the first step of immunogen preparation, the second step of coating antigen preparation, the third step of monoclonal antibody preparation, and the fourth step of sample pretreatment and detection. According to the kit, detection time is short, the average recovery rate is high, sample pretreatment is easy, on-site operation and detection can be carried out, the application range is wide, and detection cost is low; meanwhile, the kit has the advantages of being high in detection specificity and sensitivity, small in in-batch and inter-batch difference, easy and fast to operate, especially suitable for large-batch sample detection and the like.
Description
Technical field
The invention belongs to field of biological detection, specifically, is related to a kind of time resolved fluoro-immunoassay test kit of detection dodine.
Background technology
Dodine is mainly as low toxicity antibacterial, preventing and treating fruit tree, vegetable disease, predominantly effect protection, rather than without interior suction, bactericidal mechanism is the permeability for destroying cell, causes the outer of cell inclusion to blend cell death.The various main mycete disease prevented and treated on fruit tree, vegetable, nut, ornamental plant and lined tree of dodine.Therefore periodically dodine residual is carried out detecting the necessary means of the monitoring for becoming many countries.
Dodine retention analysiss generally use gas chromatography（GC）, high performance liquid chromatography（HPLC）And gas chromatography combined with mass spectrometry technology（GC/MS）, these methods are sensitive, accurate, can determine multi-medicament simultaneously, but need expensive instrument, sample pre-treatments are complicated, loaded down with trivial details time-consuming, testing cost is higher, and need professional to operate, it is difficult to meet that the needs of scene, batch, quick detection be carried out to sample.Therefore, develop a kind of simple and quick analysis method suitable for pesticide residues on-site supervision to have important practical significance.
And Timed resolved fluoroimmunoassay（TR-FIA）As its high specificity, sensitivity are high, simple to operate, cheap, and it is valued by the people the advantages of be particularly suitable for the detection of batch samples and increasingly and adopts.There is presently no the patent and document report of the time fluoroimmunoassay detected for dodine.
The content of the invention
To solve above technical problem, it is an object of the invention to provide the detection time resolved fluorometric immunoassay kitss that dodine is remained in a kind of vegetable and fruit.
The second object of the present invention is to provide a kind of detection method of the time resolved fluoro-immunoassay test kit for quickly and easily detecting dodine in vegetable and fruit, for either quantitatively or qualitatively detecting dodine residual quantity in crops.
What one of the object of the invention was realized in：The time resolved fluoro-immunoassay test kit of detection dodine, which it is critical only that being coated with plate, buffer, dodine standard solution, the antibody dried frozen aquatic productses of anti-dodine, the rabbit-anti murine antibody of europium labelling, cleaning mixture and enhancing liquid by porous is constituted.
What the two of the object of the invention were realized in：The detection method of the time resolved fluoro-immunoassay test kit of detection dodine, including the preparation and sample pre-treatments and detection of immunogen, coating antigen and monoclonal antibody, which it is critical only that：
(1) immunogenic preparation：By hapten dodine and bovine serum albumin（BSA）It is coupled, obtains immunogen（Dodine-BSA）；
(2) preparation of coating antigen：By hapten dodine and ovoserum albumin（OVA）It is coupled, obtains coating antigen（Dodine-OVA）；
(3) preparation of monoclonal antibody：
A. with the immunogen of step (1)（Dodine-BSA）Immune mouse, by hybridoma technology, the hybridoma cell strain of the monoclonal antibody for obtaining secreting anti-dodine；
B. antibody is prepared in a large number to induce ascites method in vivo, carry out purification using Protein G posts, obtain monoclonal antibody IgG of anti-dodine；
C. with 96 hole of the coating primordial covering coating plate of step (2)；
(4) pre-treatment and detection of sample：
Take and be coated with coating antigen（Dodine-OVA）Porous coating plate, add the dodine of 50 L in respective micropore, plus the anti-dodine antibody that 50 L dilutes with buffer, 25 DEG C～37 DEG C vibrate 0.5 ~ 1 hour, and cleaning mixture is washed 3 times, in addition buffer dilute 100
µL Eu3+- rabbit-anti murine antibody, 25 DEG C～37 DEG C vibrate 0.5 ~ 1 hour, and cleaning mixture is washed 6 times, plus 200
L strengthens measurement fluorescence intensity cps after liquid vibrates 5 minutes, calculates the dodine content in sample according to standard curve.
Above-mentioned solid phase carrier is porous coating plate, is coated with plate as solid phase carrier using the porous in 96 holes.
The present invention is mainly using time-resolved fluorescence immunoassay method detecting dodine.Mainly there are two aspects using the technology of Timed resolved fluoroimmunoassay：First, prepared by monoclonal antibody specific, with the immunogen immune mice being coupled, by hybridoma technology, the hybridoma cell strain of the monoclonal antibody for obtaining secreting anti-dodine；Antibody is prepared in a large number to induce ascites method in vivo, using Protein
G posts carry out purification, obtain monoclonal antibody IgG of anti-dodine.Second, Eu3+The preparation of traget antibody.
Assay method of the present invention：The basis of measure is labelled immune reaction.It is coated with the porous coating plate of dodine-OVA, addition test sample is in respective micropore, add anti-dodine antibody, oscillating reactionss, free dodine competes anti-dodine antibody with the dodine-OVA on microwell plate, cleaning mixture is washed, and does not have the dodine antibody for connecting to be removed in washing step.Add Eu3+- rabbit-anti murine antibody, is marked immunoreation, then is washed with cleaning mixture, does not have the Eu for connecting after reaction3+- rabbit-anti murine antibody is removed in washing step.Plus after strengthening liquid vibration, launch very strong fluorescence under the exciting of uviol lamp, determine its fluorescence intensity cps with time-resolved fluorescence instrument, fluorescence intensity is inversely proportional to the concentration in sample, and reference standard curve can determine that the amount of dodine in sample.
Detection method does not need expensive instrument, and sample pre-treatments are simple, energy execute-in-place detection, are widely used, and the method is sensitive, accurate, quick, easy to operate, high specificity, it is adaptable to the quick detection of gross sample.
Description of the drawings
Fig. 1 is the TR-FIA standard suppression curve figure of the dodine standard substance with antibody of the present invention.
Specific embodiment
Embodiment
1st, prepared by immunogen and coating antigen
Immunogen of the present invention（Dodine-BSA）Synthesis：Accurately weigh dodine 324mg and be dissolved in 2mL
In DMF, γ-aminobutyric acid solution under stirring, is added dropwise over, stirring reaction 3 hours adjusts reactant liquor pH 10 or so.Precipitate is removed in centrifugation.Above-mentioned reaction is added dropwise in BSA solution（320mg
BSA is dissolved in 5mL normal saline）, add N-hydroxy-succinamide（NHS）23mg, N, N- dicyclohexylcarbodiimide（DCC）45.4mg, 4 DEG C of reactions overnight, are centrifuged off precipitation, take supernatant phosphate buffer（PBS）Dialysis 3 days, changed dialysis solution, products therefrom lyophilized is saved backup in -20 DEG C per 6 hours；
Coating antigen（Dodine-OVA）Synthesis：In above-mentioned reaction, BSA is changed into after OVA, obtain reacting conjugate dodine-OVA, the conjugate is used as coating antigen when detecting as TR-FIA.
2nd, prepared by monoclonal antibody
2.1 animal immune
Immune 6 week old female Balb/c mices are distinguished with immunogen prepared by step 1, the immunizing dose of every mice is 100 g/0.2mL.First immunisation, uses aseptic 0.01mol/L
PH7.4 PBS lytic immunities are former（Dodine-BSA）, then mix with equivalent Freund's complete adjuvant, complete emulsifying, 2～3 points of injections of strength dorsal sc point；Booster immunization, uses 0.01mol/L
PH7.4 PBS lytic immunities original is mixed with equivalent Freund's complete adjuvant, fully emulsified, mouse peritoneal injection.Per minor tick 14～21 days, the 3rd time it is immune after start within 7～10 days, to immune mouse tail vein blood, to collect serum, detect mice serum potency with ELISA.It is spaced after final immunization more than 4 weeks, 3～4 days before cell fusion, 00 g/0.2mL/ of lumbar injection dodine-BSA antigen 1s only, after injection notes observation daily, it is ensured that before fusion, mice is in good condition.
It is prepared by 2.2 monoclonal antibodies
The splenocyte of separating immune mice, and carry out homogenate and prepare immune spleen cell.1 immune Balb/c mice is taken, and serum is separated as negative serum from eye socket blood-letting, is put to death.Mice carries out overall disinfection with 75% alcohol-pickled 5min.Mice extremity are fixed, then mice hypogastric region skin is clamped with tweezers, is cut off an osculum, then tear skin with tweezers, expose peritoneum, transducer set tweezers and shears, on abdominal part central authorities peritoneum cut off an osculum with shears.Transducer set tweezers and shears, cut off peritoneum with shears, expose spleen, then transducer set apparatus tweezers clamp spleen, break spleen adventitia with shears, are then placed in the homogenizer of prior sterilizing.Plus appropriate basal medium（RPMI-1640）In homogenizer, it is ground, squeezes out splenocyte, takes out the homogenate rod of homogenizer, then add appropriate basal medium（RPMI-1640）, 2min is stood, after upper cell liquid is drawn, is put in peritoneal macrophage centrifuge tube, repeat aforesaid operations 1 time.1200r/min is centrifuged 10min, removes supernatant.By 108Individual immune spleen cell and 1～2 × 107Individual SP2/0 myeloma cell is according to 1:10 or 1:During 5 ratio adds centrifuge tube, mixed, then in 1500r/min horizontal centrifugal 10min, supernatant discarded.Centrifuge tube is tipped upside down in the absorbent paper of sterilizing, liquid in pipe is blotted.Ttom of pipe is gently tapped with finger or desktop, is allowed the cell of precipitation to loosen, then centrifuge tube is placed in 37 DEG C of water-baths.It is slow by 50% in 1min
PEG 0.8mL are instilled in centrifuge tube, and side edged gently stirs sedimentation cell with pipette tip.After being further continued for stirring 30s, 1min is stood, being then slowly added into carries out the 40mL basal mediums of 37 DEG C of pre-temperatures in advance（RPMI-1640）.Plus basal medium method is：Dropwise instill dropwise to instill in 1mL, 2min in 2mL, 3min in 1min and dropwise instill 3mL, 4mL is instilled dropwise in 4min, need to be slowly added to when culture medium is added every time, and lightly stir culture base, finally remaining RPMI-1640 culture medium is slowly added into.1000r/min is centrifuged 5min, removes supernatant.Then suspended the cell for mixing with HAT culture medium, adds raising splenocyte.Appropriate HAT culture medium, mix homogeneously being added as needed, then the cell fusion drop containing feeder cells being added in 96 porocyte culture plates, dripping quantity is about 150
L/ holes.Culture plate is placed in into 37 DEG C, 5% CO2In saturated humidity incubator, cultivated.With the indirect ELISA screening positive cell clone set up.The hole of strong positive colony growth is selected, is cloned with limiting dilution assay.And to other positive holes, 24 hole amplification culture are carried out, and the supernatant in amplification culture hole being detected with indirect ELISA and indirect competitive ELISA, the cell that positive hole is to indirect ELISA and indirect competitive ELISA carries out liquid nitrogen freezing preservation.By fusion detection, and hybridoma cell strain is obtained after carrying out 3 sub-clones.Hybridoma cell strain is through repeatedly passing on, frozen, recovery, and hybridoma secretory antibody is stable.The counting of hybridoma chromosome is carried out, 20 cells will be randomly choosed per strain of hybridoma, and be carried out the counting of cell chromosome bar number, then the meansigma methodss for calculating cell chromosome bar number.Mouse boosting cell Chromosome number is 40, and the chromosome number average of SP2/0 cells is 62 ~ 68, and 20 strain of hybridoma chromosome numbers of this test acquisition are all between 92 ~ 103, average out to 96.8.Chromosome number of this hybridoma chromosome number higher than two parental cells, explanation is the hybrid product of two kinds of cells.The culture supernatant of cell strain cell secretion is taken, 1 is carried out:After 10 dilutions, antibody subtype is determined with sandwich ELISA method, the antibody subtype of the cell strain secretion is IgG1.Purification is carried out to mouse ascites using caprylic acid-ammonium.The monoclonal antibody can be used for preparation time resolved fluorometric detection kit.
The purification of 2.3 monoclonal antibodies
Purification is carried out to mouse ascites using caprylic acid-ammonium：Mouse ascites 10mL is taken, isopyknic barbitol buffer solution, appropriate silicon dioxide mixing, shaken at room temperature 30min is added.After being stored at room temperature 15min, supernatant is taken in clean centrifuge tube, 4 DEG C, 1800r/min is centrifuged 20min；Supernatant 18mL is taken, 36mL is added
0.06mol/L sodium-acetate buffers, adjust pH value to 4.5 with HCl, are sufficiently stirred for down octanoic acid 297 is slowly added in 30min
µL；Continue stirring 10min, then proceed to 4 DEG C of refrigerators and stand 2h, 4 DEG C, 15000r/min is centrifuged 30min, and supernatant volume Jing after 0.45 m membrane filtrations is 50mL；Add 5mL
The phosphate buffer of 0.1mol/L, adjusts pH value to 7.6 with NaOH, is slowly added into ammonium sulfate to final concentration of 0.277g/mL under stirring；After 4 DEG C of refrigerators stand 2h, 4 DEG C, 12000r/min centrifugation 30min abandon supernatant；Precipitation uses 5mL
The phosphate buffer of 0.1mol/L is resuspended, loads bag filter, after fully being dialysed with 5000mL 0.01mol/L pH7.2 PBSs, then is dialysed with 2000 mL distilled water, finally boils off ionized water dialysis with 3000mL tri-；Then 4 DEG C, 12000r/min centrifugation 30min abandon precipitation, collect supernatant, survey protein concentration.SDS-PAGE electrophoresis is done, the purity of monoclonal antibody is identified.
The preparation of 2.4 rabbit anti-mouse igg antibody
With Balb/C mouse IgG immune health new zealand white rabbits, the rabbit anti-mouse igg hyper-immune serum of high-titer is prepared, serum is slightly carried using saturated ammonium sulphate method, highly purified rabbit anti-mouse igg is obtained Jing after G-200 crosses post.
3.1st, reagent preparation box and detection sample
Take and be dissolved in 50 mmol/L PBS
The 5g/L rabbit-anti 1 ~ 2mL of murine antibody of pH7.0, the conversion buffered condition of Jing PD-10 posts, eluent is containing 0.155mmol/L
The 50 mmol/L Na of NaCl2CO3-NaHCO3 PH8.5 buffer.Protein peak is collected, the analysis of Jing uv absorption is quantitative（1.46A280-0.74A260）, rabbit-anti murine antibody is diluted to 2g/L with above-mentioned eluent.Take 500 ~ 1000
Rabbit-anti murine antibody after μ L dilutions adds the Eu containing 0.2 ~ 0.4mg3+-N2- [p- Carbimide .s-benzyl]-diethylenetriamine tetraacethyl（Eu3+-DTTA）Bottle in, 30 DEG C of magnetic agitation are reacted 20 hours.Reactant liquor Jing 80mmol/L
The Sepharose CL-6B posts of Tris-HCl pH7.8 buffer balance（1×40cm）Chromatography, A280Protein peak is collected in monitoring, and dilution subpackage is standby.
It is prepared by 3.2 coating plate solid phase antigens
By dodine-OVA 50mmol/LNa2CO3-NaHCO3
PH9.6 buffer is diluted to the coating buffer of 1mg/L, and coating plate each hole in 96 holes adds 100 μ L, and 4 DEG C stand overnight.Coating buffer is discarded, is flushed three times, plus 150 μ L contain 3g/L
The above-mentioned buffer blind of OVA, 4 DEG C stand overnight.Confining liquid is discarded, vacuum is drained, lath seals rearmounted -20 DEG C of freezen protectives.
The preparation of 3.3 reagents
（1）Dodine standard solution is prepared：By dodine standard substance, diluting becomes 0ng/mL, 0.01ng/mL, 0.025ng/mL, 0.1ng/mL, 0.25ng/mL, 1ng/mL, 2.5ng/mL, 10ng/mL, 25ng/mL, 100ng/mL series concentration, and diluent is 0.1mol/L
PH7.5 phosphate buffers；
（2）Buffer：8mmol/L
NaCl, 0.2% OVA, 50 μm of ol/L diethylene triamine pentacetic acid (DTPA)s（DTPA）、0.1mL/L
Tweeen-80 and 0.1%NaN350mmol/L
Tris-HCl pH7.8；
（3）Cleaning mixture is：14.5mmol/L
NaCl、0.2mL/L
Tweeen-80 and 0.2%NaN350mmol/L
Tris-HCl pH7.8；
（4）Strengthen the preparation of liquid：Added in pH3.2 Potassium Hydrogen Phthalate buffer by 15 μm of ol β-naphthoyltrifluoroacetones, 50 μm of ol TOPOs and 1mL triton x-100s, then be settled to 1L and be formulated.
The reagent that 3.4 test kits are provided
Based on the reagent of above-mentioned preparation, the present invention includes following material for the time resolved fluoro-immunoassay test kit for detecting dodine：
(1) 96 hole elisa Plates × 1 piece；
(2) dodine standard substance 1mg/mL/ bottles；
(3) anti-dodine antibody dried frozen aquatic productses, used time distill water dissolution with 0.5 mL；
(4)Eu3+- rabbit-anti murine antibody dried frozen aquatic productses, used time distill water dissolution with 0.5 mL；
(5) strengthen liquid：15mL；
(6) 10 × cleaning mixture：30mL；
(7) buffer：30 mL.
Points for attention before 3.5 measure：
A. all reagents are returned before use and is warmed to room temperature (18-30 DEG C)；
B. all reagents are put back to into 2-8 DEG C immediately after use；
If the C. big suggestion of sample size uses Multi-channel liquid transfer device；
D. during all constant-temperature incubations, it is to avoid light irradiates, and uses cap covers micropore；
E. the microwell plate and framework that need to use quantity is taken out, no microwell plate is put in former Fresco Bag and resealed together with the desiccant for providing, 2-8 DEG C is stored in.
3.6 concrete detecting steps are as follows：
Take dodine-OVA laths, add the dodine of 50 L in respective micropore, plus the anti-dodine antibody that 50 L dilutes with buffer, 25 DEG C～37 DEG C vibrate 0.5 ~ 1 hour, and cleaning mixture is washed 3 times, in addition buffer dilute 100
µL Eu3+- rabbit-anti murine antibody, 25 DEG C～37 DEG C vibrate 0.5 ~ 1 hour, and cleaning mixture is washed 6 times, plus 200
L strengthens measurement fluorescence intensity cps after liquid vibrates 5 minutes, calculates the dodine content in sample from standard curve.
3.7 follow these steps to reagent preparation box and detection Fructus Mali pumilae, Semen Maydiss, vegetable sample：
(1) the same embodiment of reagent preparation box；
(2) concrete detecting step is as follows：
Taking dodine-OVA laths, the dodine of 50 L being added in respective micropore, plus the anti-dodine antibody that 50 L dilutes with buffer, 25 DEG C～37 DEG C vibrate 0.5 ~ 1 hour, and cleaning mixture is washed 3 times, 100 L that in addition buffer dilutes
Eu3+- rabbit-anti murine antibody, 25 DEG C～37 DEG C vibrate 0.5 ~ 1 hour, and cleaning mixture is washed 6 times, plus 200
L strengthens measurement fluorescence intensity cps after liquid vibrates 5 minutes, calculates the dodine content in sample according to standard curve.
Claims (6)
1. it is a kind of detection dodine time resolved fluoro-immunoassay test kit, it is characterised in that：Plate is coated with by porous, buffer, dodine standard substance, the antibody dried frozen aquatic productses of dodine, the sheep anti-mouse antibody of europium labelling, cleaning mixture are constituted with liquid is strengthened.
2. a kind of time resolved fluoro-immunoassay test kit for detecting dodine according to claim 1, including preparation and the sample pre-treatments of immunogen, coating antigen and monoclonal antibody, it is characterised in that：
(1) dodine is coupled with bovine serum albumin, obtains immunogen；
(2) by dodine and ovoserum albumen coupling, obtain coating antigen；
(3) with the immunogen immune mice of step (1), by hybridoma technology, the hybridoma cell strain of the monoclonal antibody for obtaining secreting anti-dodine；
(4) antibody is prepared in a large number to induce ascites method in vivo, carry out purification using Protein G posts, obtain the monoclonal antibody of anti-dodine
(5) with the coating primordial covering solid phase carrier of step (2)；
(6) animal tissue is first passed through after acidolysis extraction, after MAX column purifications, is eventually adding derivative reagent and catalyst is processed, obtain product to be measured；
(7) thing to be checked of step (6) is measured into fluorescence intensity cps, reference standard curve calculates the dodine dodine content in sample.
3. the time fluoroimmunoassay test kit of dodine is detected according to claim 1, it is characterised in that：Described solid phase carrier is porous coating plate, is coated with plate as solid phase carrier using many micropores in 96 holes.
4. the time fluorescence resolved immuno analytic process test kit of dodine is detected according to claim 1, it is characterised in that：Described derivative reagent is butylamine.
5. the Timed resolved fluoroimmunoassay test kit of dodine is detected according to claim 1, it is characterised in that：Described catalyst is itrile group diethyl phosphate.
6. the time fluoroimmunoassay test kit of dodine is detected according to claim 1, it is characterised in that：The step (6) and (7) are specially and take the micropore for being coated with dodine-OVA coating plate, the sample that 50 L are handled well is added in respective micropore, the dodine antibody for adding 50 L and being diluted with buffer, 25 ~ 37 DEG C vibrate 0.5 ~ 1 hour, cleaning mixture is washed three times, in addition 100 L Eu of buffer dilution3+- sheep anti-mouse antibody, 25 ~ 37 DEG C vibrate 0.5 ~ 1 hour, and cleaning mixture is washed six times, plus 200 L strengthen measurement fluorescence intensity cps after liquid vibrates 5 minutes, calculate the dodine content in sample from standard curve.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN110208235A (en) * | 2019-06-26 | 2019-09-06 | 贵州大学 | The fluorescence probe and its detection method of pesticide dodine in a kind of detection water |
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| CN1598586A (en) * | 2004-09-07 | 2005-03-23 | 江苏省农业科学院 | Immune antibody for detecting pesticide organic phosphorus residus and its application |
| US20080199972A1 (en) * | 2005-05-02 | 2008-08-21 | Frank Sellrie | Spectroscopic Method For the Detection of Analytes |
| CN101776601A (en) * | 2010-01-25 | 2010-07-14 | 泸州老窖集团有限责任公司 | Method for fast detecting pesticide residue |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110208235A (en) * | 2019-06-26 | 2019-09-06 | 贵州大学 | The fluorescence probe and its detection method of pesticide dodine in a kind of detection water |
| CN110208235B (en) * | 2019-06-26 | 2022-02-01 | 贵州大学 | Fluorescent probe for detecting pesticide dodine in water and detection method thereof |
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Application publication date: 20170426 |