CN105588940A - Preparation of time-resolved fluorescence immunoassay kit for plasticizer detection - Google Patents
Preparation of time-resolved fluorescence immunoassay kit for plasticizer detection Download PDFInfo
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Abstract
The present invention discloses a time-resolved fluorescence immunoassay kit for plasticizer detection, wherein the time-resolved fluorescence immunoassay kit comprises a porous coated plate, a buffer liquid, a plasticizer standard substance, a plasticizer antibody freeze-drying substance, europium labeled goat anti-mouse antibody, a washing liquid and an enhancing liquid. The detection method comprises: (1) immunogen preparation; (2) coated antigen preparation; (3) monoclonal antibody preparation; and (4) sample pre-treatment and detection. According to the present invention, the time-resolved fluorescence immunoassay kit has advantages of short detection time, high average recovery rate, simple sample pre-treatment, on-site operation detection, wide application, low detection cost, strong detection specificity, low intra-assay or inter-assay difference, high sensitivity and simple and rapid operation, and the like, and is particularly suitable for detection of a large number of samples.
Description
Technical field
The invention belongs to field of biological detection, specifically, relate to a kind of preparation of the time-resolved fluoroimmunoassay kit that detects plasticiser.
Background technology
Plasticiser is a kind of flexibility of material or additive of material liquefaction of increasing. Phthalate plasticiser is classified as doubtful Environmental Hormone, its bio-toxicity mainly belongs to estrogen and antiandrogen activity, can cause endocrinopathy, infringement organism Reproductive Performance, comprise that reproduction rate reduces, miscarriage, born defect, abnormal sperm count, damage of testis, also can cause malignant tumour, cause deformed child, cause children's sexual precocity.
Dibutyl phthalate (Dibutylphthalate, DBP) is a kind of conventional plasticiser, also, as the additive of adhesive and printing-ink, also can be used as a kind of ectoparasiticide. It has stronger genotoxicity, can cause that male fecundity declines, especially larger to children's toxicity. DBP is mainly present in artificial leather, plastic products, cosmetics, essence and agricultural chemicals.
At present, the method for detection plasticiser mainly contains high performance liquid chromatography HPLC, gas chromatography GC etc. Wherein, high performance liquid chromatography HPLC, gas chromatography GC are quantitative detection methods, and accurately and reliably, shortcoming is that detection limit is higher, instrument and equipment is comparatively expensive to result, and complicated Sample pretreatment method, has limited the extensive use of the method. Enzyme-linked Immunosorbent Assay ELISA method is a kind of accurate, reliable, quick, special detection method, is suitable for the rapid screening of gross sample, in recent years extensive use.
The present invention is intended to set up a kind of time-resolved fluoroimmunoassay (TR-FIA) kit that detects plasticiser (DBP), due to its high specificity, highly sensitive, simple to operate, cheap, and be particularly suitable for the advantages such as the detection of batch samples and be more and more valued by the people and adopt. Patent and the bibliographical information of the time fluoroimmunoassay detecting for plasticiser are at present less.
Summary of the invention
For solving above technical problem, the object of the present invention is to provide plasticiser in a kind of white wine (DBP) to differentiate fluorescence immunoassay kit residual detection time.
Two of object of the present invention is to provide one to detect quickly and easily the detection method of the time resolved fluoro-immunoassay kit of plasticiser in white wine (DBP), for quantitatively or qualitatively detecting white wine plasticiser (DBP) residual quantity.
One of the object of the invention is achieved in that the time resolved fluoro-immunoassay kit that detects plasticiser (DBP), and its key is to be made up of the coated plate of porous, buffer solution, plasticiser standard solution, the antibody dried frozen aquatic products of anti-plasticiser, rabbit anti-mouse antibody, cleaning solution and the enhancing liquid of europium mark.
Two of the object of the invention is achieved in that the detection method of the time resolved fluoro-immunoassay kit that detects plasticiser (DBP), comprise preparation and sample pre-treatments and the detection of immunogene, coating antigen and monoclonal antibody, its key is:
(1) immunogenic preparation: by haptens plasticiser (DBP) and bovine serum albumin(BSA) (BSA) coupling, obtain immunogene (DBP-BSA);
(2) preparation of coating antigen: by haptens plasticiser and ovoserum albumin (OVA) coupling, obtain coating antigen (DBP-OVA);
(3) preparation of monoclonal antibody:
A. immunogene (DBP-BSA) immune mouse of using step (1), by hybridoma technology, obtains the hybridoma cell strain of the monoclonal antibody of secreting anti-plasticiser;
B. to induce a large amount of Dispersal risks of ascites method in body, use ProteinG post to carry out purifying, obtain the monoclonal antibody IgG of anti-plasticiser;
C. use the coated plate in coated 96 holes of coating antigen of step (2);
(4) pre-treatment of sample and detection:
Get 5mL sample in clean teat glass, add the chromatographically pure n-hexane of 2mL, the 3min that vibrates after closing the lid, then leaves standstill, and gets supernatant liquor 1mL after layering, dries up in clean glassware room temperature nitrogen, dries up the rear methyl alcohol with 1mL35% and redissolves rear to be measured;
Preparation 35% methanol solution: get 35mL methyl alcohol and add in 65ml deionized water and mix;
Get the coated plate of porous that is coated with coating antigen (DBP-OVA), add standard items/sample of 50 μ L in micropore separately, add the anti-plasticiser antibody of 50 μ L with buffer solution dilution, 25 DEG C～37 DEG C vibrate 0.5 ~ 1 hour, cleaning solution is washed 3 times, in addition 100 μ LEu of buffer solution dilution3+-rabbit anti-mouse antibody, 25 DEG C～37 DEG C vibrate 0.5 ~ 1 hour, and cleaning solution is washed 6 times, adds 200 μ L enhancing liquid vibrations and measures fluorescence intensity cps after 5 minutes, according to the content of plasticizing agent in calibration curve calculation sample.
Above-mentioned solid phase carrier is the coated plate of porous, adopts the coated plate of porous in 96 holes as solid phase carrier.
The present invention mainly adopts time-resolved fluorescence immunoassay method to detect plasticiser. Adopt the technology of time resolved fluoro-immunoassay method to mainly contain two aspects: the first, monoclonal antibody specific preparation, with the immunogen immune mouse of coupling, by hybridoma technology, obtains the hybridoma cell strain of the monoclonal antibody of secreting anti-plasticiser; To induce a large amount of Dispersal risks of ascites method in body, use ProteinG post to carry out purifying, obtain the monoclonal antibody IgG of anti-plasticiser. The second, Eu3+The preparation of labelled antibody.
Assay method of the present invention: the basis of mensuration is labelled immune reaction. Be coated with the coated plate of porous of DBP-OVA, add test sample in micropore separately, add again anti-plasticiser antibody, oscillating reactions, DBP-OVA on free plasticiser and microwell plate competes anti-plasticiser antibody, cleaning solution washing, does not have the plasticiser antibody connecting to be removed in washing step. Add Eu3+-rabbit anti-mouse antibody, carries out labelled immune reaction, then with cleaning solution washing, the Eu not connecting after reaction3+-rabbit anti-mouse antibody is removed in washing step. Add and strengthen after liquid vibration, the very strong fluorescence of transmitting under the exciting of uviol lamp, measures its fluorescence intensity cps with time resolution luminoscope, and the concentration in fluorescence intensity and sample is inversely proportional to, and reference standard curve can be determined the amount of plasticiser in sample.
Detection method of the present invention does not need expensive instrument, and sample pre-treatments is simple, can detect by execute-in-place, is widely used, and the method is sensitive, accurate, quick, and easy and simple to handle, high specificity is applicable to the fast detecting of gross sample.
Brief description of the drawings
Fig. 1 is that the TR-FIA standard of plasticiser standard items of the present invention and antibody suppresses curve map.
Detailed description of the invention
Embodiment
1, immunogene and coating antigen preparation
Synthesizing of immunogene of the present invention (DBP-BSA): accurately take plasticiser 324mg and be dissolved in 2mLN, in dinethylformamide, dropwise add GABA solution under stirring, stirring reaction 3 hours, regulates reactant liquor pH10 left and right. The centrifugal sediment of removing. Above-mentioned reaction is dropwise added to (320mgBSA is dissolved in 5mL physiological saline) in BSA solution, add again N-hydroxy-succinamide (NHS) 23mg, N, N-dicyclohexylcarbodiimide (DCC) 45.4mg, 4 DEG C of reactions are spent the night, the centrifugal precipitation of removing, get phosphate buffer for supernatant (PBS) dialysis 3 days, within every 6 hours, change dislysate, by products therefrom lyophilized, save backup in-20 DEG C.
Synthesizing of coating antigen (DBP-OVA): in above-mentioned reaction, BSA is changed into after OVA, obtain reacting conjugate DBP-OVA, this conjugate uses as coating antigen while detection as TR-FIA.
2, monoclonal antibody preparation
2.1 animal immune
The immunogene of preparing by step 1 female Balb/c mouse in respectively immune 6 week age, the immunizing dose of every mouse is 100 μ g/0.2mL. First immunisation, with aseptic 0.01mol/LpH7.4PBS lytic immunity former (DBP-BSA), then mixes with equivalent Freund's complete adjuvant, emulsification completely, the injection of subcutaneous point 2～3, strength back; Booster immunization, mixes with equivalent Freund's complete adjuvant with 0.01mol/LpH7.4PBS lytic immunity is former, fully emulsified, mouse peritoneal injection. Every minor tick 14～21 days, starts immune mouse tail vein blood after the 3rd immunity for 7～10 days, collects serum, detects mice serum tire with ELISA. After last immunity, interval is more than 4 weeks, and before Fusion of Cells 3～4 days, lumbar injection DBP-BSA antigen 1 00 μ g/0.2mL/ only, after injection, noted observing by every day, and before ensureing to merge, mouse is in good condition.
2.2 monoclonal antibody preparations
The splenocyte of separating immune mouse, and carry out homogenate and prepare immune spleen cell. Get 1 immune Balb/c mouse,, put to death as negative serum from eye socket bloodletting separation of serum. 75% alcohol-pickled 5min for mouse, carries out overall disinfection. Mouse four limbs are fixed, then clamped mouse lower abdomen skin with tweezers, cut off an osculum, then tear skin with tweezers, expose peritonaeum, transducer set tweezers and scissors are cut off an osculum with scissors on belly central authorities peritonaeum. Transducer set tweezers and scissors, cut off peritonaeum with scissors, exposes spleen, then transducer set apparatus clamps spleen with tweezers, spleen adventitia broken with scissors, then puts into the homogenizer of prior sterilizing. Add appropriate basal medium (RPMI-1640) in homogenizer, grind, squeeze out splenocyte, take out the homogenate rod of homogenizer, add again appropriate basal medium (RPMI-1640), leave standstill 2min, after upper strata cell liquid is drawn, put into peritoneal macrophage centrifuge tube, repeat aforesaid operations 1 time. The centrifugal 10min of 1200r/min, removes supernatant. By 108Individual immune spleen cell and 1～2 × 107Individual SP2/0 myeloma cell adds in centrifuge tube according to the ratio of 1:10 or 1:5, mixes, then in 1500r/min horizontal centrifugal 10min, supernatant discarded. Centrifuge tube is tipped upside down on the blotting paper of sterilizing, liquid in pipe is blotted. Knock gently and manage at the end with finger or desktop, make the cell of precipitation loosening, then centrifuge tube is placed in to 37 DEG C of water-baths. In 1min, slowly 50%PEG0.8mL is splashed in centrifuge tube, limit edged stirs sedimentation cell with pipette tip gently. Continue again to stir after 30s, leave standstill 1min, then slowly add the 40mL basal medium (RPMI-1640) that carries out in advance 37 DEG C of pre-temperature. Adding basal medium method is: in 1min, dropwise splash into 1mL, in 2min, dropwise splash into 2mL, in 3min, dropwise splash into 3mL, in 4min, dropwise splash into 4mL, in the time adding culture medium, need slowly add at every turn, and stir culture base lightly, finally remaining RPMI-1640 culture medium is slowly added. The centrifugal 5min of 1000r/min, removes supernatant. Then the cell mixing with the suspension of HAT culture medium, then add raising splenocyte. Add as required appropriate HAT culture medium, mix, then the Fusion of Cells drop that contains feeder cells is added on 96 porocyte culture plates, dripping quantity is about 150 μ L/ holes. Culture plate is placed in to 37 DEG C, 5%CO2In saturated humidity incubator, cultivate. With the indirect ELISA screening positive cell clone of setting up. Select the hole of strong positive colony growth, clone with limiting dilution assay. And to other positive holes, carry out 24 holes and expand and cultivate, the supernatant of expansion culture hole is detected with indirect ELISA and indirect competitive ELISA, to indirect ELISA and indirect competitive ELISA all the cell in positive hole carry out liquid nitrogen frozen preservation. By fusion detection, and carry out obtaining hybridoma cell strain after 3 subclones. Hybridoma cell strain through repeatedly going down to posterity, frozen, recovery, hybridoma secretory antibody is stable. Carry out the chromosomal counting of hybridoma, by 20 cells of the random selection of every strain of hybridoma, carry out the counting of cell chromosome number, then calculate the mean value of cell chromosome number. Mouse boosting cell chromosome number is 40, and the chromosome number average of SP2/0 cell is 62 ~ 68, and the 20 strain of hybridoma chromosome numbers that this test obtains are all between 92 ~ 103,96.8 of average out to. This hybridoma chromosome number is higher than the chromosome number of two parental cells, and explanation is the hybridization product of two kinds of cells. Get the culture supernatant of cell line emiocytosis, carry out after 1:10 dilution, measure antibody subtype with sandwich ELISA method, the antibody subtype of this cell line secretion is IgG1. Adopt caprylic acid-ammonium to carry out purifying to mouse ascites. This monoclonal antibody can be used for preparation time and differentiates fluorescence detection reagent kit.
The purifying of 2.3 monoclonal antibodies
Adopt caprylic acid-ammonium to carry out purifying to mouse ascites: get mouse ascites 10mL, add isopyknic barbitol buffer solution, appropriate silica mixes, room temperature vibration 30min. Room temperature leaves standstill after 15min, gets supernatant in clean centrifuge tube, and 4 DEG C, the centrifugal 20min of 1800r/min; Get supernatant 18mL, add 36mL0.06mol/L sodium-acetate buffer, with HCl adjust pH to 4.5, under fully stirring, in 30min, slowly add sad 297 μ L; Continue to stir 10min, then proceed to 4 DEG C of refrigerators and leave standstill 2h, 4 DEG C, the centrifugal 30min of 15000r/min, supernatant volume after 0.45 μ m membrane filtration is 50mL; Add the phosphate buffer of 5mL0.1mol/L, with NaOH adjust pH to 7.6, under stirring, slowly adding ammonium sulfate to final concentration is 0.277g/mL; 4 DEG C of refrigerators leave standstill after 2h, and 4 DEG C, the centrifugal 30min of 12000r/min, abandons supernatant; Precipitation is resuspended with the phosphate buffer of 5mL0.1mol/L, packs bag filter into, after fully dialysing, then uses 2000mL distill water dialysis with 5000mL0.01mol/LpH7.2PBS buffer solution, finally boils off ionized water dialysis with 3000mL tri-; Then 4 DEG C, the centrifugal 30min of 12000r/min, abandons precipitation, collects supernatant, surveys protein concentration. Do SDS-PAGE electrophoresis, the purity of qualification monoclonal antibody.
The preparation of 2.4 rabbit anti-mouse igg antibody
With Balb/C mouse IgG immune health new zealand white rabbit, prepare the rabbit anti-mouse igg hyper-immune serum of high-titer, adopt saturated ammonium sulphate method slightly to carry serum, after crossing post, G-200 obtains highly purified rabbit anti-mouse igg.
3, sample pretreating method
Get 5mL Wine Sample in clean teat glass, add the chromatographically pure n-hexane of 2mL, 3min vibrates after closing the lid, then leave standstill, after layering, get supernatant liquor 1mL, dry up in clean glassware room temperature nitrogen, dry up the rear methyl alcohol with 1mL35% and redissolve rear to be measured;
Preparation 35% methanol solution: get 35mL methyl alcohol and add in 65ml deionized water and mix.
4, prepare kit and detect sample
Get the 5g/L rabbit anti-mouse antibody 1 ~ 2mL that is dissolved in 50mmol/LPBSpH7.0, through the conversion buffered condition of PD-10 post, eluent is the 50mmol/LNa containing 0.155mmol/LNaCl2CO3-NaHCO3PH8.5 buffer solution. Collect protein peak, through quantitatively (1.46A280-0.74A260) of UV absorption analysis, dilute rabbit anti-mouse antibody to 2g/L with above-mentioned eluent. The rabbit anti-mouse antibody of getting after 500 ~ 1000 μ L dilutions adds the Eu containing 0.2 ~ 0.4mg3+-N2-[p-isocyanic acid-benzyl]-diethylenetriamine tetraacethyl (Eu3+-DTTA) bottle in, 30 DEG C of magnetic agitation reaction 20 hours. Reactant liquor is through SepharoseCL-6B post (1 × 40cm) chromatography by 80mmol/LTris-HClpH7.8 buffer solution balance, A280Protein peak is collected in monitoring, and dilution packing is for subsequent use.
4.2 coated plate solid phase antigen preparations
By DBP-OVA 50mmol/LNa2CO3-NaHCO3PH9.6 buffer solution is diluted to the coating buffer of 1mg/L, and the each hole of coated plate, 96 holes adds 100 μ L, and 4 DEG C of placements are spent the night. Discard coating buffer, rinse three times, add the above-mentioned buffer solution sealing of 150 μ L containing 3g/LOVA, 4 DEG C of placements are spent the night. Discard confining liquid, vacuum is drained, rearmounted-20 DEG C of freezing preservations of lath sealing.
The preparation of 4.3 reagent
(1) plasticiser standard solution preparation: by plasticiser standard items, dilution becomes 0ng/mL, 0.01ng/mL, 0.03ng/mL, 0.09ng/mL, 0.27ng/mL, 0.81ng/mL, 2.43ng/mL, 7.29ng/mL, 25ng/mL series concentration, dilution is 0.1mol/LpH7.5 phosphate buffer;
(2) buffer solution: 8mmol/LNaCl, 0.2%OVA, 50 μ mol/L diethylene triamine pentacetic acid (DTPA)s (DTPA), 0.1mL/LTweeen-80 and 0.1%NaN350mmol/LTris-HClpH7.8;
(3) cleaning solution is: 14.5mmol/LNaCl, 0.2mL/LTweeen-80 and 0.2%NaN350mmol/LTris-HClpH7.8.
(4) preparation of enhancing liquid: added in pH3.2 Potassium Hydrogen Phthalate buffer solution by 15 μ mol β-naphthoyltrifluoroacetones, 50 μ mol TOPOs and 1mL triton x-100, then it is formulated to be settled to 1L.
The reagent that 4.4 kits provide
Based on the reagent of above-mentioned preparation, the present invention comprises following material for detection of the time resolved fluoro-immunoassay kit of plasticiser:
(1) 96 ELISA Plate × 1 piece, hole;
(2) plasticiser standard items 1mg/mL/ bottle;
(3) anti-plasticiser antibody dried frozen aquatic products, the used time dissolves with 0.5mL distilled water;
(4)Eu3+-rabbit anti-mouse antibody dried frozen aquatic products, the used time dissolves with 0.5mL distilled water;
(5) strengthen liquid: 15mL;
(6) 10 × cleaning solutions: 30mL;
(7) buffer solution: 30mL;
4.5 measure points for attention before:
A. before using, all reagent is gone up to room temperature (18-30 DEG C).
B. after using, immediately all reagent is put back to 2-8 DEG C.
If C. the large suggestion of sample size is used Multi-channel liquid transfer device.
D. in all constant-temperature incubation processes, avoid light to irradiate, use cap covers micropore.
E. take out the microwell plate and the framework that need by quantity, no microwell plate is put in former Fresco Bag and together with the drier providing and resealed, be stored in 2-8 DEG C.
4.6 concrete detecting steps are as follows:
Get DBP-OVA lath, add standard items/sample of 50 μ L in micropore separately, add the anti-plasticiser antibody of 50 μ L with buffer solution dilution, 25 DEG C～37 DEG C vibrations 0.5 ~ 1 hour, cleaning solution is washed 3 times, in addition 100 μ LEu of buffer solution dilution3+-rabbit anti-mouse antibody, 25 DEG C～37 DEG C vibrate 0.5 ~ 1 hour, and cleaning solution is washed 6 times, adds 200 μ L enhancing liquid vibrations and measures fluorescence intensity cps, the content of plasticizing agent from calibration curve calculation sample after 5 minutes.
4.7 follow these steps to prepare kit and detect Wine Sample:
(1) prepare kit with 4 of embodiment;
(2) concrete detecting step is as follows:
Get 5mL Wine Sample in clean teat glass, add the chromatographically pure n-hexane of 2mL, 3min vibrates after closing the lid, then leave standstill, after layering, get supernatant liquor 1mL, dry up in clean glassware room temperature nitrogen, dry up the rear methyl alcohol with 1mL35% and redissolve rear to be measured;
Preparation 35% methanol solution: get 35mL methyl alcohol and add in 65ml deionized water and mix;
Get DBP-OVA lath, add standard items/sample of 50 μ L in micropore separately, add the anti-plasticiser antibody of 50 μ L with buffer solution dilution, 25 DEG C～37 DEG C vibrations 0.5 ~ 1 hour, cleaning solution is washed 3 times, in addition 100 μ LEu of buffer solution dilution3+-rabbit anti-mouse antibody, 25 DEG C～37 DEG C vibrate 0.5 ~ 1 hour, and cleaning solution is washed 6 times, adds 200 μ L enhancing liquid vibrations and measures fluorescence intensity cps after 5 minutes, according to the content of plasticizing agent in calibration curve calculation sample.
Claims (6)
1. a time resolved fluoro-immunoassay kit that detects plasticiser, is characterized in that: by the coated plate of porous, and buffer solution, plasticiser standard items, the antibody dried frozen aquatic products of plasticiser, the sheep anti-mouse antibody of europium mark, cleaning solution and enhancing liquid form.
2. detect according to claim 1 a detection method for the time resolved fluoro-immunoassay kit of plasticiser, comprise preparation and the sample pre-treatments of immunogene, coating antigen and monoclonal antibody, it is characterized in that:
(1) by plasticiser and bovine serum albumin(BSA) coupling, obtain immunogene;
(2) by plasticiser and ovoserum albumen coupling, obtain coating antigen;
(3) with the immunogen immune mouse of step (1), by hybridoma technology, obtain the hybridoma cell strain of the monoclonal antibody of secreting anti-plasticiser;
(4) to induce a large amount of Dispersal risks of ascites method in body, use ProteinG post to carry out purifying, obtain the monoclonal antibody of anti-plasticiser
(5) be coated with solid phase carrier with the coating antigen of step (2);
(6) by animal tissue first through acidolysis extract after, after MAX column purification, finally add derivative reagent and catalyst to process, obtain product to be measured;
(7) thing to be checked of step (6) is measured to fluorescence intensity cps, the content of plasticizing agent in reference standard curve calculation sample.
3. the detection method that detects according to claim 1 the time fluoroimmunoassay kit of plasticiser, is characterized in that: described solid phase carrier is the coated plate of porous, adopts the coated plate of many micropores in 96 holes as solid phase carrier.
4. the time fluorescence that detects according to claim 1 plasticiser is differentiated the detection method of immunoassay kit, it is characterized in that: described derivative reagent is butylamine.
5. the detection method that detects according to claim 1 the time resolved fluoro-immunoassay method kit of plasticiser, is characterized in that: described catalyst is itrile group diethyl phosphate.
6. detect according to claim 1 the detection method of the time fluoroimmunoassay kit of plasticiser, it is characterized in that: described step (6) and (7) are specially gets the coated plate of micropore that is coated with plasticiser-OVA, add sample that 50 μ L handle well in micropore separately, add the plasticiser antibody of 50 μ L with buffer solution dilution, 25 ~ 37 DEG C vibrate 0.5 ~ 1 hour, cleaning solution is washed three times, in addition 100 μ LEu of buffer solution dilution3+-sheep anti-mouse antibody, 25 ~ 37 DEG C vibrate 0.5 ~ 1 hour, and cleaning solution is washed six times, adds 200 μ L enhancing liquid vibrations and measures fluorescence intensity cps, the content of plasticizing agent from calibration curve calculation sample after 5 minutes.
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| CN109752529A (en) * | 2017-11-06 | 2019-05-14 | 镇江华维检测技术有限公司 | Detect the preparation of the time-resolved fluoroimmunoassay kit of plasticiser |
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| CN108226465A (en) * | 2016-12-15 | 2018-06-29 | 江苏维赛科技生物发展有限公司 | Detect the time-resolved fluoroimmunoassay kit of aflatoxin B1 |
| CN108226504A (en) * | 2016-12-15 | 2018-06-29 | 江苏维赛科技生物发展有限公司 | Detect the time-resolved fluoroimmunoassay kit of sodium sulfocyanate |
| CN109752522A (en) * | 2017-11-05 | 2019-05-14 | 镇江华维检测技术有限公司 | Detect the time-resolved fluoroimmunoassay kit and its detection method of o-phenyl phenol in fruits and vegetables |
| CN109752554A (en) * | 2017-11-05 | 2019-05-14 | 江苏维赛科技生物发展有限公司 | The time-resolved fluoroimmunoassay kit of Aflatoxins M1 in a kind of milk |
| CN109752529A (en) * | 2017-11-06 | 2019-05-14 | 镇江华维检测技术有限公司 | Detect the preparation of the time-resolved fluoroimmunoassay kit of plasticiser |
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