CN107315091A - One kind detection Cistofuran metabolite quantum dot immune chromatographic test paper, preparation method and applications - Google Patents
One kind detection Cistofuran metabolite quantum dot immune chromatographic test paper, preparation method and applications Download PDFInfo
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Abstract
Cistofuran metabolite quantum dot immune chromatographic test paper is detected the invention provides one kind, the test paper is pasted onto on support plate and constituted successively by sample pad, microporous fiber membranes, reaction film, adsorptive pads, has Cistofuran metabolite specific antibody quantum dot probe on the microporous fiber membranes.Quantum dot fluorescence immuno-chromatographic test paper strip detection method has the advantages that sensitivity height, high specificity, cost are low, simple to operate, detection time is short, is adapted to various units uses, storage is simple, long shelf-life, qualitative, quantitative all may be used.Wherein, using the Cistofuran metabolite monoclonal antibody specific of high specific, it is ensured that the reliability of testing result；It is easy, quick, accurate with the method for ELISA test strip Cistofuran metabolite of the present invention.
Description
Technical field
The present invention relates to field of immunodetection, and in particular to one kind detection Cistofuran metabolite quantum dot immune chromatography examination
Paper, preparation method and applications.
Technical background
Furantoin (Nitrofurantion), belongs to Nitrofuran antibiotics, is a kind of broad-spectrum antibiotics, has
The characteristic of good antibacterial action and medicine power, medicine is to most of gram-positive bacterias, Gram-negative bacteria, fungi and protozoon
There is killing action etc. pathogen, because price is relatively low and effect is good, once widely used in culture fishery.But study for a long period of time
Prove, Nitrofuran metabolites and metabolin can make experimental animal occur canceration, gene mutation and have teratogenesis tire side effect,
The multinational use for having prohibited such medicine in treatment and feed.European Union provided against and used itrofurans comprehensively nineteen ninety-five
Antibacterials, the limit value of Nitrofuran metabolites and metabolite residue, which must not be, in animal derived food detects.U.S. in 2002
State completely forbids is used for food-borne animal by Nitrofuran metabolites.Food and drug administration (FDA) announcement in 2004
Forbid the 11 kinds of list of substance used in import animal derived food, including nitrofurazone and furazolidone.China
The Ministry of Agriculture issues No. 193 bulletins in April, 2002, discloses《Food animal disables veterinary drug and other compound inventories》, regulation
Nitrofuran antibiotics are prohibitted the use of in all food animals, are again included aquatic products Nitrofuran metatolites within 2003
Remain Supervisory Surveillance Program.
The most popular method for being used for detecting Cistofuran metabolite at present is high performance liquid chromatography (HPLC), LC-MS
Method (LC-MS, LC-MS/MS) and ELISA etc..LC-MS, LC-MS/MS and HPLC have high sensitivity high specific
Feature, but sample pre-treatments are cumbersome, reagent consumption big, detection time length, equipment are expensive, need professional and technical personnel to be grasped
Make, be not suitable for the screening of progress field quick detection and a large amount of samples, it is difficult to the need for the examination for meeting great amount of samples.It is enzyme-linked to exempt from
Although the sensitivity of epidemic disease analytic approach is high, high specificity needs the instrument and equipments such as ELIASA, and detection time is long, and requires ability
Domain professional and technical personnel operates.These methods are difficult to the quick detection and Site Detection of batch samples.
Quantum dot (Quantum dots, QDs), also known as semiconductor nano, are that one kind is launched under light source activation
The semiconductor nano crystal grain of bright fluorescence, is mainly made up of II~VI race or III~group Ⅴ element, such as cadmium telluride (CdTe), selenium
Cadmium (CdSe) etc..Compared with conventional fluorescent material, QDs advantage has：With quantum effect and good photism；Have
Wider excitation wavelength and narrower launch wavelength；Stability is good；With good bio-compatibility；Fluorescence lifetime is longer.By
There is above good characteristic in quantum dot, make it possible to turn into the powerful of Molecular Detection and medical diagnosis, more than nearly 20
The development in year, progressively to clinical detection, disease since the application studies such as cell, tissue mark's imaging, Cellular tracking, living imaging
The field directions such as sick control, food safety detection are developed.
A kind of simple and fast that quantum dot mark immunity-chromatography technology is set up on the basis of immunity percolation technology it is immune
Detection technique is learned, is marked thing using fluorescence quantum, quantum dot and antibody are passed through into chemically react (COOH and NH2) be coupled at
Together, when antibody is combined with antigentic specificity, quantum dot is just marked on determined antigen, is presented not under the irradiation of uviol lamp
Same fluorescence intensity.Wherein, the presence or absence of control line (C lines) color decides the validity of test strips, and detection line (T lines) has
Nothing then represents the presence or absence of object.In addition to the plurality of advantages for possessing ELISA, the one of ELISA is also overcomed
It is a little not enough.The technical operation is simple and quick, and reaction generally only needs several minutes to dozens of minutes, and testing result is with macroscopic aobvious
Vitta brings interpretation, and safety non-pollution is with a wide range of applications.
The content of the invention
The present invention in view of the shortcomings of the prior art and defect, utilizes quantum dot multi-wavelength excitation, high intensity fluorescent emission, hair
Penetrating the fluorescent characteristic that peak is narrow, peak shape is symmetrical, stability of photoluminescence is good, there is provided a kind of quantum of new detection Cistofuran metabolite
The preparation method and applications of point immuno-chromatographic test paper strip, according to the photoluminescent property of quantum dot and Cistofuran metabolite concentration it
Between relation, metabolin is detected in test strips, the quantitative detection of quantum dot ELISA test strip metabolin is realized, it is existing to solve
Sensitivity is low, complex operation, the problems such as equipment requirement is high.
Cistofuran metabolite quantum dot immune chromatographic test paper is detected the invention provides one kind, the test paper is by sample
Pad, microporous fiber membranes, reaction film, adsorptive pads are pasted onto on support plate and constituted successively, have furans appropriate on the microporous fiber membranes
Because of metabolin specific antibody-quantum dot probe.
Cistofuran metabolite specific antibody in the Cistofuran metabolite specific antibody-quantum dot probe is
Prepared using Cistofuran metabolite hapten-carrier protein conjugate as immunogene, the Cistofuran metabolite is special
Heterogenetic antibody is Cistofuran metabolite monoclonal antibody specific.
It is provided with the reaction film and the detection being made is coated with by Cistofuran metabolite hapten-carrier protein conjugate
Line and the control line being made up of rabbit-anti mouse antiantibody coating.
A kind of preparation method for detecting Cistofuran metabolite quantum dot immune chromatographic test paper, comprises the following steps：
1) the Cistofuran metabolite specific antibody of preparation is added in the quantum dot of preparation, obtains furantoin generation
Thank to thing specific antibody-quantum dot probe；Cistofuran metabolite specific antibody-quantum dot probe is uniformly sprayed in micropore
Glass fibre membrane surface obtains microporous fiber membranes；
2) Cistofuran metabolite hapten-carrier protein conjugate and rabbit-anti mouse antiantibody are sprayed onto cellulose nitrate respectively
It is corresponding on plain film to detect on line position and control line position and be coated with, obtain reaction film；
3) by microporous fiber membranes, reaction film and sample pad, adsorptive pads, support plate are fitted together, and load cartridge, plus dry
Drying prescription is sealed, and obtains detecting the preparation method of Cistofuran metabolite quantum dot immune chromatographic test paper.
5. the preparation method of detection Cistofuran metabolite quantum dot immune chromatographic test paper according to claim 4,
It is characterized in that：The preparation method of the Cistofuran metabolite specific antibody comprises the following steps：
1) Cistofuran metabolite and p -carboxybenzaldehyde are reacted, prepares Cistofuran metabolite haptens；
2) by Cistofuran metabolite haptens and carrier protein couplet, Cistofuran metabolite hapten-carrier is prepared
Protein conjugate；
3) mouse immuning ball protein G immune healths mountain rabbit is extracted, rabbit-anti mouse antiantibody is obtained；With Cistofuran metabolite half
Mouse is immunized in antigen-carrier protein conjugate, by mouse boosting cell and murine myeloma cell by fusion, screening, is divided
The hybridoma cell strain of Cistofuran metabolite monoclonal antibody specific is secreted, Cistofuran metabolite specific monoclonal is obtained
Antibody, is used as Cistofuran metabolite specific antibody.
The Cistofuran metabolite hapten-carrier protein conjugate is by Cistofuran metabolite haptens and carrier
Albumen coupling is obtained, and the Furaxone metabolite haptens is reacted by Cistofuran metabolite and p -carboxybenzaldehyde
Arrive, the carrier protein can be bovine serum albumin(BSA).
The preparation method of the quantum dot is：
By CdCl2.2.5H2O is dissolved in the water, and leads to nitrogen deoxygenation 20-30mim, and it is jointly molten to add TGA, uses
The regulation of NaOH solution is 11.0 ± 0.2 to pH value, leads to high pure nitrogen protection, adds Na2TeO3Solution and NaBH4, continue to stir
Lower heated solution is to seething with excitement, and reaction backflow obtains red CdTe QDs as quantum dot.
The quantum dot immune chromatograph test strip of detection Cistofuran metabolite described above is in detection animal derived food
The application of middle Nitrofuran metatolites metabolin.
The material that the support plate can not absorb water for PVC support plates or other hard；The sample pad can for suction strainer paper or
Filter paper for oil；The adsorptive pads are blotting paper；The reaction film can be nitrocellulose filter；There is test strips card outside the test strips
Shell, the epivalve that gets stuck, which corresponds at sample pad location, is provided with well, and well sees sample pad；The epivalve correspondence that gets stuck
Observation window, observation window visible detection line and control line are provided with reaction film location.
It is metabolized it is a further object to provide furantoin in the above-mentioned ELISA test strip animal tissue of one kind application
Thing is remained, and it includes step：
(1) Sample pretreatment；
(2) detected with test strips；
(3) testing result is analyzed.
The Cleaning Principle (as shown in Figure 1, 2) of test strips of the present invention：Measuring samples drop after processing is added on test strips
In well, measuring samples liquid is together with after the quantum dot probe combination in microporous fiber membranes to reaction membrane diffusion；If treating sample
The content of Cistofuran metabolite is high in product liquid, then the Cistofuran metabolite in diffusion process in measuring samples liquid can be with quantum
Point probe is combined, and then completely encloses the antigen-combining site of Cistofuran metabolite on probe, is prevented on probe and reaction film
Cistofuran metabolite hapten-carrier protein conjugate is combined, and detection line does not develop the color, and rabbit-anti mouse secondary antibody then can be with probe knot
Close, control line colour developing；If the content of Cistofuran metabolite is low in measuring samples liquid or nothing, the antigen binding site on probe
It can not be closed, and then probe can be combined with Cistofuran metabolite hapten-carrier protein-coupled antigen on reaction film, detection
Line develops the color, while rabbit-anti mouse secondary antibody can also be combined with probe, control line colour developing.If control line does not develop the color, test paper failure.
Qualitative detection or half-quantitative detection：Reacted reaction film line domain is loaded using ultra violet lamp, seen by naked eyes
Examine band to light situation, when control line (C) shows fluorescent bands, and detection line (T) is judged to the positive when not developing the color；Work as control
Line (C) shows fluorescent bands, detection line (T) while also showing that red stripes, is judged to feminine gender；When nature controlling line (C) is not shown
Go out fluorescent bands, then no matter whether detection line (T) shows fluorescent bands, and it is invalid that the test paper is judged to.
Qualitative detection：80 μ L mixed liquors are taken to be added in the well in test strips, reaction 3-10min is that knot can be read
Really；
Quantitative detection：A series of molecular criteria solution to be checked of various concentrations is equipped with, quantitative fluorescence analysis instrument is then utilized
Immunochromatography detection is carried out respectively, by doing standard curve to each peak area corresponding concentration, then unknown sample to be checked is carried out
Same processing, obtains peak area, testing molecule content in sample is learnt according to calibration curve formula.
The bright advantageous effects of we are：Quantum dot fluorescence immuno-chromatographic test paper strip detection method of the present invention and enzyme
The methods such as connection immunoabsorption, gas chromatography, liquid chromatography are compared, and with sensitivity height, high specificity, cost is low, grasp
Make that simple, detection time is short, be adapted to various units and use, storage is simple, long shelf-life, qualitative, quantitative all can the advantages of.Wherein,
Using the Cistofuran metabolite monoclonal antibody specific of high specific, it is ensured that the reliability of testing result；With the present invention
The method of ELISA test strip Cistofuran metabolite is easy, quick, accurate.
Figure of description
The schematic diagram of the quantum dot immune chromatograph test strip of Fig. 1 Cistofuran metabolites；
The quantum dot immune chromatograph test strip outline drawing of Fig. 2 Cistofuran metabolites；
The quantum dot immune chromatographic test paper testing result process decision chart of Fig. 3 Cistofuran metabolites；
The quantum dot immune chromatograph test strip quantitative measurement standard curve map of Fig. 4 Cistofuran metabolites；
Wherein, 1, support plate, 2 sample pads, 3 microporous fiber membranes, 4 detection lines, 5 control lines, 6 nitrocellulose membranes, 7 absorb
Pad, 8 quantum dot probes, 9 Cistofuran metabolites, 10 rabbit-anti mouse antiantibodys, 11 Cistofuran metabolites and carrier protein couplet
Thing, 12 wells, 13 observation windows, 14 cartridges.
Embodiment
The preparation of the test paper of the detection Cistofuran metabolite of embodiment 1
The preparation of 1 microporous fiber membranes
1.1 react Cistofuran metabolite and p -carboxybenzaldehyde, prepare Cistofuran metabolite haptens；By furans
It is appropriate because of metabolite hapten and carrier protein couplet, prepare Cistofuran metabolite hapten-carrier protein conjugate；Use furans
It is appropriate because mouse is immunized in metabolite hapten-carrier protein couplet thing, by mouse boosting cell and murine myeloma cell by fusion,
Screening, obtains secreting the hybridoma cell strain of Cistofuran metabolite monoclonal antibody specific, obtains Cistofuran metabolite
Monoclonal antibody specific (anti-NPAHD McAb).
0.120g CdCl is added in 1.2 round-bottomed flasks2.2.5H2O is dissolved in 100mL tri-distilled waters, leads to nitrogen deoxygenation 20-
30 mim, add 72.7 μ L TGAs and are dissolved in jointly in 200mL tri-distilled waters, and pH is arrived with 1.00mol/L NaOH solution regulation
It is worth for 11.0 or so, leads to high pure nitrogen 30min, Na is added under nitrogen protection2TeO3Solution 210 μ L and 20mg NaBH4, after
The lower heated solution of continuous stirring is to seething with excitement, and reaction backflow obtains red CdTe QDs, is used as quantum dot (QDs).
1.3 take the μ L of carboxyl water solubility QDs 600 prepared in 1.2, add 18 μ L carbodiimide hydrochloride (1mg/mL)
With lucifuge concussion 30min after 800 μ L methanol mixing, the beta -mercaptoethanol for adding 8 μ L is terminated, and takes 10mL anti-
NPAHD McAb (3mg/mL) are added to be mixed with the QDs of activation, lucifuge concussion 2h, is added the stable QDs of mercaptoethanol, is entered
Row dialysis.3min is centrifuged after dialysis under the conditions of 1,6300r/min, supernatant is removed, precipitation is resuspended in 10mmol/l PBS
(pH7.4) in, 4 DEG C of preservations obtain Cistofuran metabolite specific antibody-quantum dot probe.
Cistofuran metabolite specific antibody-quantum dot probe is taken out placement to room temperature by 1.4, is diluted with PBS-TBN
100-1000 times, into quantum dot-labeled probe solution, then uniformly sprays quantum dot-labeled antibody probe in glass fibre membrane table
Face, drying at room temperature, 4 DEG C of preservations.PBS-TBN used is sodium chloride nacl containing 20mM-150mM, 0.02%-0.1% tweens
Tween-20,2mg/mL-10m/mL bovine serum albumin(BSA) BSA, 0.05% sodium azide NaN310mM-100mM phosphate delay
Fliud flushing PB, obtains microporous fiber membranes.
The preparation of 2 reaction films
2.1 extract mouse immuning ball protein G (IgG) immune healths mountain rabbit, obtain rabbit-anti mouse antiantibody；
2.2 fix the middle part for being pasted onto PVC backings with nitrocellulose filter, and nitrocellulose filter is placed in into ZX1000
On point film instrument platform, Cistofuran metabolite hapten-carrier protein conjugate coating buffer solution is adjusted into concentration to 1mg/mL,
And with coating buffering, also regulation concentration is to 0.5mg/mL by rabbit-anti mouse secondary antibody, according to 1uL/cm film liquid amount, by furantoin generation
Thank to thing hapten-carrier protein conjugate, and rabbit-anti mouse antiantibody is sprayed onto on reaction film in corresponding detection line and control line
It is coated with, at intervals of 4mm between detection line and control line, 25~37 DEG C of placement oven 2 hours is placed in constant temperature and humidity guarantor
Hide standby in case, coating buffer solution used is to contain 20mM-150mM sodium chloride nacls, 0.05%-3% polyethylene glycol
PEG20000,0.2%-1% trehalose, 2mg/mL-10m/mL bovine serum albumin(BSA)s BSA, 0.05% sodium azide NaN3's
10mM-100 mM phosphate buffers PB；Obtain reaction film.
The preparation of 3 sample pads
Suction strainer paper is placed in containing 0.5% bovine serum albumin(BSA) (volumn concentration), 0.1mol/L phosphate buffers
(pH7.2) immersion 2h in, 37 DEG C of baking 2h are standby, obtain sample pad.
The preparation of 4 detection Cistofuran metabolite test paper
The test paper is made up of support plate, sample pad, microporous fiber membranes, reaction film, adsorptive pads；It is the sample pad, micro-
Hole tunica fibrosa, reaction film and adsorptive pads are pasted onto on support plate in order successively, and the end of sample pad is overlapping with microporous fiber membranes
The connected, end of microporous fiber membranes is overlapping with reaction film to be connected, and the end of reaction film is overlapping with adsorptive pads to be connected, the beginning of sample pad
End is alignd with the top of support plate, and the end of adsorptive pads is alignd with the end of support plate；There are detection line and control on the reaction film
Line processed, in the bar wire (as shown in Figure 1, 2) perpendicular with the length of the test strips；It is wide that 3mm is cut into CM4000 cutting machines
Test strips, test strips are loaded into test card, quantum dot fluorescence immuno-chromatographic test paper strip is made.
Wherein, the support plate is PVC support plates；The sample pad is suction strainer paper；The adsorptive pads are blotting paper；It is described
Reaction film is nitrocellulose filter.Above-mentioned test strips are loaded in test paper cartridge, preserved in 2~8 DEG C of environment, the term of validity 12
Month.
Embodiment 2 detects the application of Cistofuran metabolite test paper
1 Sample pretreatment
(5.0 ± 0.05) g homogeneous sample is taken into 50mL polystyrene centrifuge tubes, the trichloroacetic acids of 5mL 10% are added, then
100 μ L derivatization reagents are added, 3min is fully vibrated, 1.5h is placed at 60 DEG C；Taking-up sequentially adds 1mL 0.5mol/L phosphoric acid
The potassium solution of hydrogen two, 1.5mL 2mol/L sodium hydroxide solutions and 10mL ethyl acetate, vibrate 10s, and the high sample of fat content is light
Under micro oscillation 8~10；More than 3000g, (20~25 DEG C) centrifugation 5min of room temperature；The glass for taking 8mL ethyl acetate phases to be dried to 10mL
In glass test tube, in being dried up under 50~60 DEG C of water-bath nitrogen streams, 1mL n-hexanes are added, with vortex instrument whirling motion 30s, 0.6mL are added
0.2mol/L phosphate buffers, whirling motion 10s is fully mixed；In more than 3000g, (20~25 DEG C) centrifugation 5min of room temperature, removing
Layer organic phase, removing the μ L of layer 100 is used to analyze.
2 test strips sensitivity
100 μ L measuring samples liquid are drawn with micropipettor to add in test strips sample pad, microporous fiber membranes is fully soaked
Enter in solution；(20-25 DEG C) of room temperature is incubated after 5min, is taken out test strips, is observed its detection line by gross visualization, as a result see figure
2, its detection line fluorescence and the fluorescence of 0ng/mL blank control detection lines when Cistofuran metabolite concentration is less than 0.1ng/mL
There is no notable difference；When Cistofuran metabolite concentration is 0.3ng/mL, its detection line fluorescence is detected with 0ng/mL blank controls
The fluorescence of line substantially dies down；When Cistofuran metabolite content is more than or equal to 1ng/mL scopes, its detection line goes out without color
It is existing；The range estimation of this test strips judges that detection is limited to 1ng/mL.Instrument, which is read, with fluorescent test paper strip determines paper slip detection line (T) and control line
(C) fluorescent value, according to the fluorescent value of machine-readable each concentration Cistofuran metabolite and the ratio (B/B of 0 concentration fluorescent value0) and it is each
Standard curve is drawn in the logarithm value fitting of Cistofuran metabolite concentration, sees Fig. 3.ELISA test strip Cistofuran metabolite concentration
Scope is 0.01-100ng/mL；The good standard curve Cistofuran metabolite range of linearity is 0.1ng/mL to 10ng/mL,
The sensitivity of quantitative testing test paper bar is 0.58ng/mL.
3 test strips specificity
By Cistofuran metabolite analog (furantoin, furazolidone, furaltadone, nitrofurazone, furazolidone generation
Thank thing, AMOZ, Furacilin metabolite) it is diluted with 0.2mol/L phosphate buffer (pH7.2), use
Test strips described in embodiment 7 are detected, determine the specificity of test strips, and experimental result is shown in Table 1.From the result of table 1,
It is metabolized with the ELISA test strip furantoin, furazolidone, furaltadone, nitrofurazone, Furaxone metabolite, furaltadone
During the medicines such as thing, Furacilin metabolite, test strips control line and detection line develop the color, illustrate this test strips to these medicines without
Cross reaction, specificity is preferably.
The test strips of table 1 specificity
4 TIANZHU XINGNAO Capsuls
Cistofuran metabolite is added in the flesh of fish, crab meat and shrimp blank sample, addition be respectively 0.2ng/g,
0.5ng/g, 2ng/g, 5ng/g, carry out carrying out after sample treatment according to preceding method, with ELISA test strip, and are read with fluorescence
Instrument surveys its detection line fluorescent value, and Cistofuran metabolite content and its rate of recovery are calculated according to the standard curve of foundation.Experiment knot
Fruit is shown in Table 2.By
Table 2 understands that TIANZHU XINGNAO Capsul is 81.5%~108.2%, variation within batch coefficient in the flesh of fish, crab meat and shrimp tissue
For 8.7%~12.5%.
Table 2 is oppressed, the rate of recovery and the coefficient of variation (n=5) in crab meat and shrimp tissue
。
Claims (8)
1. one kind detection Cistofuran metabolite quantum dot immune chromatographic test paper, it is characterised in that：The test paper is by sample pad, micro-
Hole tunica fibrosa, reaction film, adsorptive pads are pasted onto on support plate and constituted successively, have furantoin metabolism on the microporous fiber membranes
Thing specific antibody-quantum dot probe.
2. detection Cistofuran metabolite quantum dot immune chromatographic test paper according to claim 1, it is characterised in that：It is described
Cistofuran metabolite specific antibody in Cistofuran metabolite specific antibody-quantum dot probe is with furantoin generation
Thank to thing hapten-carrier protein conjugate to prepare as immunogene, the Cistofuran metabolite specific antibody is furan
Mutter appropriate because of metabolin monoclonal antibody specific.
3. detection Cistofuran metabolite quantum dot immune chromatographic test paper according to claim 1, it is characterised in that：It is described
It is provided with reaction film and the detection line that is made is coated with and by rabbit-anti mouse by Cistofuran metabolite hapten-carrier protein conjugate
The control line that antiantibody coating is made.
4. a kind of preparation method for detecting Cistofuran metabolite quantum dot immune chromatographic test paper, it is characterised in that：Including following
Step：
1) the Cistofuran metabolite specific antibody of preparation is added in the quantum dot of preparation, obtains Cistofuran metabolite
Specific antibody-quantum dot probe；Cistofuran metabolite specific antibody-quantum dot probe is uniformly sprayed in micropore glass
Tunica fibrosa surface obtains microporous fiber membranes；
2) Cistofuran metabolite hapten-carrier protein conjugate and rabbit-anti mouse antiantibody are sprayed onto nitrocellulose filter respectively
It is above corresponding to detect on line position and control line position and be coated with, obtain reaction film；
3) by microporous fiber membranes, reaction film and sample pad, adsorptive pads, support plate are fitted together, and load cartridge, plus drier
Sealing, obtains detecting the preparation method of Cistofuran metabolite quantum dot immune chromatographic test paper.
5. the preparation method of detection Cistofuran metabolite quantum dot immune chromatographic test paper according to claim 4, it is special
Levy and be：The preparation method of the Cistofuran metabolite specific antibody comprises the following steps：
1) Cistofuran metabolite and p -carboxybenzaldehyde are reacted, prepares Cistofuran metabolite haptens；
2) by Cistofuran metabolite haptens and carrier protein couplet, Cistofuran metabolite hapten-carrier albumen is prepared
Conjugate；
3) mouse immuning ball protein G immune healths mountain rabbit is extracted, rabbit-anti mouse antiantibody is obtained；It is anti-with Cistofuran metabolite half
Mouse is immunized in original-carrier protein couplet thing, by mouse boosting cell and murine myeloma cell by fusion, screening, is secreted
The hybridoma cell strain of Cistofuran metabolite monoclonal antibody specific, obtains Cistofuran metabolite specific monoclonal and resists
Body, is used as Cistofuran metabolite specific antibody.
6. the preparation method of detection Cistofuran metabolite quantum dot immune chromatographic test paper according to claim 5, it is special
Levy and be：The Cistofuran metabolite hapten-carrier protein conjugate is by Cistofuran metabolite haptens and carrier
Albumen coupling is obtained, and the Furaxone metabolite haptens is reacted by Cistofuran metabolite and p -carboxybenzaldehyde
Arrive, the carrier protein can be bovine serum albumin(BSA).
7. the preparation method of detection Cistofuran metabolite quantum dot immune chromatographic test paper according to claim 4, it is special
Levy and be：The preparation method of the quantum dot is：
CdCl2.2.5H2O is dissolved in the water, leads to nitrogen deoxygenation 20-30mim, it is jointly molten to add TGA, molten with NaOH
Liquid regulation is 11.0 ± 0.2 to pH value, leads to high pure nitrogen protection, adds Na2TeO3 solution and NaBH4, continues to stir lower heat
Solution is to seething with excitement, and reaction backflow obtains red CdTe QDs as quantum dot.
8. a kind of quantum dot immune chromatograph test strip for detecting Cistofuran metabolite nitro furan in detection animal derived food
Mutter the application of metabolite metabolin.
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Cited By (4)
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| CN108226490A (en) * | 2018-01-03 | 2018-06-29 | 河南省农业科学院 | A kind of quantum dot test strips for quickly detecting micro ethiprole and preparation method thereof |
| CN109884293A (en) * | 2019-03-29 | 2019-06-14 | 中国海洋大学 | A kind of food allergen rapid detection method based on quantum dot fluorescence |
| CN112557661A (en) * | 2020-10-31 | 2021-03-26 | 上海海洋大学 | Magnetic immunochromatographic test strip and method for rapidly detecting pre-S2 antigen of hepatitis B virus |
| CN113156112A (en) * | 2021-04-25 | 2021-07-23 | 渤海大学 | Fluorescence immunochromatography kit for simultaneously detecting four nitrofuran metabolites and preparation method and application thereof |
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| CN109884293A (en) * | 2019-03-29 | 2019-06-14 | 中国海洋大学 | A kind of food allergen rapid detection method based on quantum dot fluorescence |
| CN112557661A (en) * | 2020-10-31 | 2021-03-26 | 上海海洋大学 | Magnetic immunochromatographic test strip and method for rapidly detecting pre-S2 antigen of hepatitis B virus |
| CN113156112A (en) * | 2021-04-25 | 2021-07-23 | 渤海大学 | Fluorescence immunochromatography kit for simultaneously detecting four nitrofuran metabolites and preparation method and application thereof |
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