CN107132348A - A kind of method of color fluorescence particle marker antibody and the test strips prepared using it - Google Patents
A kind of method of color fluorescence particle marker antibody and the test strips prepared using it Download PDFInfo
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- CN107132348A CN107132348A CN201710095977.XA CN201710095977A CN107132348A CN 107132348 A CN107132348 A CN 107132348A CN 201710095977 A CN201710095977 A CN 201710095977A CN 107132348 A CN107132348 A CN 107132348A
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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
- 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
- G01N33/569—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses
- G01N33/56983—Viruses
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- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
- G01N2333/005—Assays involving biological materials from specific organisms or of a specific nature from viruses
- G01N2333/08—RNA viruses
- G01N2333/115—Paramyxoviridae, e.g. parainfluenza virus
- G01N2333/13—Canine distemper virus
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
- G01N2333/005—Assays involving biological materials from specific organisms or of a specific nature from viruses
- G01N2333/08—RNA viruses
- G01N2333/15—Retroviridae, e.g. bovine leukaemia virus, feline leukaemia virus, feline leukaemia virus, human T-cell leukaemia-lymphoma virus
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Abstract
The test strips prepared the invention provides a kind of method of color fluorescence particle marker antibody and using it, the present invention is to include but is not limited to CDV, canine parvovirus, hepatitis infectiosa canis virus, canine coronavirus, hydrophobin, feline leukaemia virus, marek's disease virus, the animal virus such as NDV are object, the antibody of corresponding color fluorescence microballoon mark is prepared for chemical covalent method, applied in animal virus detection.A kind of method of color fluorescence particle marker antibody prepared by the present invention and the test strips prepared using it, it is bright colored detection lines that colour developing occurs during detection, provides qualitative results；The content for providing animal virus in sample then can be also quantified, test result is relatively sharp and stability is higher, and the long period can be preserved at room temperature.
Description
Technical field
The test strips prepared the present invention relates to a kind of method of color fluorescence particle marker antibody and using it, the method
The test strips of preparation can be used for including but not limited to CDV, canine parvovirus, hepatitis infectiosa canis virus, canine coronavirus, mad dog
Sick virus, feline leukaemia virus, marek's disease virus, the quick detection of the animal virus such as NDV, belong to medical science and examine
Test field.
Background technology
With other microbial ratios, virus structure, chemical composition, modes of reproduction and to drug susceptibility in terms of have
Significant difference.The characteristics of animal virosis has strong infectiousness, serious popular extensive, harm and the high incidence of disease, thus it is how fast
Speed has diagnosed into control and treated the key factor of these animal virosis.Currently, each veterinary laboratories detection animal virosis
Main to utilize serological test and molecular biology test, these methods are often time-consuming longer, do not have the work of quick detection
With.
In recent years, immune chromatography method is increasingly ripe, and it has time-consuming short, easy to operate, with low cost suitable for scene
The features such as great amount of samples is quickly tested, achieves significant progress in the detection field of animal virus.Wherein, collaurum because
The advantages such as easy preparation, good biocompatibility, color-variable become mark raw material the most frequently used in immunochromatographic assays.
In animal virus prison detection, collaurum can realize qualitative detection easily because of its strikingly color, and be examined by photoelectric sensor
Absorption and scattering of the colloid gold particle to light are surveyed, quantitative detection can be achieved.But, the production cost of body gold is of a relatively high, uses
Physical absorption makes its requirement to raw materials such as antibody and coating antigens higher；When gold grain is excessive, label is again unstable to lead
Cause differences between batches big；And collaurum signal is weaker, sensitivity is low to limit its application.Therefore need it is a kind of cheap,
Easily prepare, stable, the high marking particle of sensitivity, which substitutes collaurum, is used for the immunochromatography that animal virus is detected.
The content of the invention
Based on this, object of the present invention is to provide a kind of method of color fluorescence particle marker antibody and it is used
The test strips of preparation realize the quick detection of animal virus, and solve collaurum cost height, and signal is weak, and p-wire colour developing is not clear
It is aobvious, quantitatively detect the problems such as differences between batches are big.Test strips prepared by the method for color fluorescence particle marker antibody, can be simultaneously real
The qualitative and quantitative detection of existing highly sensitive animal virus.
The color fluorescence particle target antibody that the present invention is provided, to be bonded europium3+Color fluorescence latex beads exemplified by system
Preparation Method, comprises the following steps：
(1) quantitative color fluorescence latex beads solution is added in cushioning liquid and diluted, ultrasonic 1.5min obtains coloured silk
Color fluorescent microsphere dispersion liquid.
(2) it is rapid in the color fluorescence latex beads dispersion liquid into step (1) to instill activator, it is quickly placed into vortex
On oscillator after low speed concussion 1min, it is fixed on turbula shaker, at room temperature concussion activation 30min.
(3) after activation terminates, by the color fluorescence latex beads dispersion liquid in step (2) under 26 DEG C, 14000 rpm,
Supernatant is removed after centrifugation 12min, is cleaned, after being centrifuged again with the same terms, discarded with the cushioning liquid in step (1)
Clear liquid, is redissolved with cushioning liquid in step (4) and is resuspended.
(4) monoclonal antibody of accordingly dynamic thing virus is taken, cushioning liquid is added and is diluted to 0.085mg/mL.
(5) nanoparticulate dispersion redissolved in step (3) is added in the antibody diluent in step (4), at once
1.5min is shaken on vortex oscillator, subsequent ultrasound 1.5min is then attached on turbula shaker and shakes anti-at room temperature
Answer 1h.
(6) after reaction terminates, by solution ultrasound 1min in step (5), at 26 DEG C, under 14000rpm, after centrifugation 12min
Supernatant is removed, confining liquid is added, is equally then attached on turbula shaker concussion reaction 1h at room temperature.
(7) confining liquid cleaning will be added after solution centrifugal in step (6) again, at 26 DEG C, under 14000rpm, centrifugation
Supernatant is removed after 12min, the antibody of color fluorescence microballoon mark is can obtain.
The cushioning liquid of step (1) is 2- (N- morpholines) ethyl sulfonic acid cushioning liquid, and its pH is 7.2.
Activator is EDC＆NHS activators in step (2).
The cushioning liquid and the cushioning liquid of step (4) redissolved in step (3) is boric acid acid buffering solution, its pH
It is worth for 7.8.
Present invention also offers a kind of test strips of the antibody of the color fluorescence microballoon mark prepared based on the above method,
Characterized in that, the structure of kit is as shown in Figure 1：Sample pad (2), one end of sample pad are pasted in plastic bottom board (1) one end
Close crimping is coated with the pad (3) of the animal virus antibody of color fluorescence microballoon mark, and pad (3) one end is closely pressed
Connect and detection line T1 (5) and nature controlling line C (6) are coated with nitrocellulose NC films (4), NC films, the other end connection water suction of NC films
Pad (7) and form test paper, test paper loads plastics and got stuck interior formation test card.Including following number of assembling steps：
(1) re-suspension liquid is added in the color fluorescence microballoon of antibody labeling, blown and beaten repeatedly, subsequent ultrasound 5min is until precipitation
It is completely dissolved.
(2) the color fluorescence microspheres solution of the antibody labeling in step (1) is sprayed at pad with film metal spraying machine is drawn,
Then 1h is dried at 37 DEG C in an oven.
(3) control line and detection line are wanted into coated antibody, respectively with after suitable diluted, with a stroke film metal spraying
It is sprayed on nitrocellulose filter by machine respectively, and 1h is dried in an oven at 37 DEG C.
(4) pad, sample pad are pasted in assembling, and sealing is stayed overnight, and are allowed to fully bonding.
The solubility of the antibody-solutions of colored fluorescent microsphere mark is 1.5mg/mL in step (2), and coated weight is 5 μ L/cm.
The coated antibody of control line of nitrocellulose filter is anti-mouse lgG polyclonal antibodies in step (3), and coated weight is
0.9μL/cm；The coated antibody of detection line is animal virus monoclonal antibody, and coated weight is 0.9 μ L/cm.
The invention provides the application method of this animal virus test strip：
(1) pipettor is used, takes the animal blood serum, blood plasma, whole blood sample of determined volume to be added in sample diluting liquid,
After being sufficiently mixed, take the test fluid of determined volume to be added in the loading slot of test strips, wait in 3~6min, if only Quality Control
There are color stripes in line, and it is negative to show test result, and corresponding animal virus is free of in sample；If control line and p-wire
Color stripes are shown, test result is the positive, is continued waiting for 15min, is examined using supporting fluorescent quantitative detector device
Survey, the content for providing animal virus in sample can be quantified.
The invention provides application of the test strips of above-mentioned preparation method preparation in animal virus detection.
Relative to prior art, the examination that the present invention is proposed the method for color fluorescence particle marker antibody and prepared using it
Paper slip, there is following advantage：
(1) method of antibody labeling of the present invention is the functional group using color fluorescence microsphere surface, using change
Learn mode and the antibody binding of covalent bonding so that the color fluorescence microballoon of antibody labeling is more stable compared with collaurum.
(2) when test paper of the present invention is used for animal virus detection, qualitative detection result is can obtain in 3~6min,
Supporting fluorescent quantitative detector device is then used in 15min, you can quantitatively obtain accurate result, it is qualitative fixed to realize
Amount is detected simultaneously.
(3) when test paper of the present invention is used for animal virus detection, minimum detection limit can reach 4ng/mL, and sensitivity is high
In colloidal gold immunity chromatography.
Brief description of the drawings
The illustrated embodiment that the accompanying drawing of the present invention is used for providing in a further understanding of the present invention, the present invention is used to say
The bright present invention, does not constitute to the present invention and limits.
Fig. 1 is the structural representation of the test strip of colored fluorescent microsphere mark in present invention.
The test strips for the CDV Antibody preparation that Fig. 2 marks for the color fluorescence microballoon in the embodiment of the present invention one
Test result.
Embodiment
The present invention is described in detail below, but is not limited to the scope of the present invention.
Embodiment one
A kind of test strips of the CDV Antibody preparation of color fluorescence microballoon mark, comprise the following steps：
(1) quantitative color fluorescence latex beads solution is added in cushioning liquid and diluted, ultrasonic 1.5min obtains coloured silk
Color fluorescent microsphere dispersion liquid.
(2) it is rapid in the color fluorescence latex beads dispersion liquid into step (1) to instill activator, it is quickly placed into vortex
On oscillator after low speed concussion 1min, it is fixed on turbula shaker, at room temperature concussion activation 30min.
(3) after activation terminates, by the color fluorescence latex beads dispersion liquid in step (2) under 26 DEG C, 14000 rpm,
Supernatant is removed after centrifugation 12min, is cleaned, after being centrifuged again with the same terms, discarded with the cushioning liquid in step (1)
Clear liquid, is redissolved with cushioning liquid in step (4) and is resuspended.
(4) monoclonal antibody of CDV is taken respectively, is separately added into cushioning liquid and is diluted to 0.085 mg/mL.
(5) nanoparticulate dispersion redissolved in step (3) is added in the antibody diluent in step (4), at once
1.5min is shaken on vortex oscillator, subsequent ultrasound 1.5min is then attached on turbula shaker and shakes anti-at room temperature
Answer 1h.
(6) after reaction terminates, by solution ultrasound 1min in step (5), at 26 DEG C, under 14000rpm, after centrifugation 12min
Supernatant is removed, confining liquid is added, is equally then attached on turbula shaker concussion reaction 1h at room temperature.
(7) confining liquid cleaning will be added after solution centrifugal in step (6) again, at 26 DEG C, under 14000rpm, centrifugation
Supernatant is removed after 12min, the antibody of color fluorescence microballoon mark is can obtain.
The cushioning liquid of step (1) is 2- (N- morpholines) ethyl sulfonic acid cushioning liquid, and its pH is 7.2.
Activator is EDC＆NHS activators in step (2).
The cushioning liquid and the cushioning liquid of step (4) redissolved in step (3) is boric acid acid buffering solution, its pH
It is worth for 7.8.
Present invention also offers a kind of test strips of the antibody of the color fluorescence microballoon mark prepared based on the above method,
Including following number of assembling steps：
(1) re-suspension liquid is added in the color fluorescence microballoon of antibody labeling, blown and beaten repeatedly, subsequent ultrasound 5min is until precipitation
It is completely dissolved.
(2) the color fluorescence microspheres solution of the antibody labeling in step (1) is sprayed at pad with film metal spraying machine is drawn,
Then 1h is dried at 37 DEG C in an oven.
(3) control line and detection line are wanted into coated antibody, respectively with after suitable diluted, with a stroke film metal spraying
It is sprayed on nitrocellulose filter by machine respectively, and 1h is dried in an oven at 37 DEG C.
(4) pad, sample pad are pasted in assembling, and sealing is stayed overnight, and are allowed to fully bonding, be can obtain canine distemper and the white blood of cat
Sick Viral diagnosis test card.
The solubility of the antibody-solutions of colored fluorescent microsphere mark is 1.5mg/mL in step (2), and coated weight is 5 μ L/cm.
The coated antibody of control line of nitrocellulose filter is anti-mouse lgG polyclonal antibodies in step (3), and coated weight is
0.9μL/cm；The coated antibody of detection line is CDV monoclonal antibody, and coated weight is 0.9 μ L/cm.
The invention provides the application method of this animal virus test strip：
(1) pipettor is used, takes the dog serum, blood plasma, whole blood sample of determined volume to be added in sample diluting liquid, fills
Divide after mixing, take the test fluid of determined volume to be added in the loading slot of test strips, wait in 3~6min, if only nature controlling line
There are color stripes, it is CDV in feminine gender, sample to show test result；Behave excellently if control line and p-wire show
Vitta band, test result is the positive, is continued waiting for 15min, is detected using supporting fluorescent quantitative detector device, can quantify and give
Go out the content of animal virus in sample.
Experiment and the result
1. the test strips of the CDV Antibody preparation marked using present case color fluorescence microballoon carry out hundstaupe pyreticosis
Poison test.As shown in Fig. 2 negative and positive all two groups of parallel testings of carry out amount, two groups of tests succeed.After sample-adding during 5min,
Negative sample testing result only has control line colored lines occur, and two colored lines occurs in positive testing result,
Detected during 15min using supporting fluorescent quantitative detector device, two groups of positive quantitative results are respectively 18.6ng/mL, 19.1ng/
mL。
Embodiment two
A kind of test strips of the feline leukaemia virus Antibody preparation of color fluorescence microballoon mark, comprise the following steps：
(8) quantitative color fluorescence latex beads solution is added in cushioning liquid and diluted, ultrasonic 1.5min obtains coloured silk
Color fluorescent microsphere dispersion liquid.
(9) it is rapid in the color fluorescence latex beads dispersion liquid into step (1) to instill activator, it is quickly placed into vortex
On oscillator after low speed concussion 1min, it is fixed on turbula shaker, at room temperature concussion activation 30min.
(10) after activation terminates, by the color fluorescence latex beads dispersion liquid in step (2) at 26 DEG C, under 14000rpm,
Supernatant is removed after centrifugation 12min, is cleaned, after being centrifuged again with the same terms, discarded with the cushioning liquid in step (1)
Clear liquid, is redissolved with cushioning liquid in step (4) and is resuspended.
(11) monoclonal antibody of feline leukaemia virus is taken respectively, is separately added into cushioning liquid and is diluted to 0.085 mg/mL.
(12) nanoparticulate dispersion redissolved in step (3) is added in the antibody diluent in step (4), stood
It is engraved on vortex oscillator and shakes 1.5min, subsequent ultrasound 1.5min is then attached on turbula shaker and shaken at room temperature
React 1h.
(13) after reaction terminates, by solution ultrasound 1min in step (5), at 26 DEG C, under 14000rpm, after centrifugation 12min
Supernatant is removed, confining liquid is added, is equally then attached on turbula shaker concussion reaction 1h at room temperature.
(14) confining liquid cleaning will be added after solution centrifugal in step (6) again, at 26 DEG C, under 14000rpm, centrifugation
Supernatant is removed after 12min, the antibody of color fluorescence microballoon mark is can obtain.
The cushioning liquid of step (1) is 2- (N- morpholines) ethyl sulfonic acid cushioning liquid, and its pH is 7.2.
Activator is EDC＆NHS activators in step (2).
The cushioning liquid and the cushioning liquid of step (4) redissolved in step (3) is boric acid acid buffering solution, its pH
It is worth for 7.8.
Present invention also offers a kind of test strips of the antibody of the color fluorescence microballoon mark prepared based on the above method,
Including following number of assembling steps：
(5) re-suspension liquid is added in the color fluorescence microballoon of antibody labeling, blown and beaten repeatedly, subsequent ultrasound 5min is until precipitation
It is completely dissolved.
(6) the color fluorescence microspheres solution of the antibody labeling in step (1) is sprayed at pad with film metal spraying machine is drawn,
Then 1h is dried at 37 DEG C in an oven.
(7) control line and detection line are wanted into coated antibody, respectively with after suitable diluted, with a stroke film metal spraying
It is sprayed on nitrocellulose filter by machine respectively, and 1h is dried in an oven at 37 DEG C.
(8) pad, sample pad are pasted in assembling, and sealing is stayed overnight, and are allowed to fully bonding, be can obtain canine distemper and the white blood of cat
Sick Viral diagnosis test card.
The solubility of the antibody-solutions of colored fluorescent microsphere mark is 1.5mg/mL in step (2), and coated weight is 5 μ L/cm.
The coated antibody of control line of nitrocellulose filter is anti-mouse lgG polyclonal antibodies in step (3), and coated weight is
0.9μL/cm；The coated antibody of detection line is feline leukaemia virus monoclonal antibody, and coated weight is 0.9 μ L/cm.
The invention provides the application method of this feline leukaemia virus test strip：
(1) pipettor is used, takes cat serum, blood plasma, the whole blood sample of determined volume to be added in sample diluting liquid, fills
Divide after mixing, take the test fluid of determined volume to be added in the loading slot of test strips, wait in 3~6min, if only nature controlling line
There are color stripes, it is feline leukaemia virus in feminine gender, sample to show test result；If control line and p-wire are shown
Color stripes, test result is the positive, is continued waiting for 15min, is detected, can quantified using supporting fluorescent quantitative detector device
Provide in sample
The content of animal virus.
Experiment and the result
1. the test strips of the feline leukaemia virus Antibody preparation marked using present case color fluorescence microballoon are also with the white blood of cat
50 parts of clinical samples are determined exemplified by virus, the coincidence rate of interval result of determination and sample value is 94.2%.It is of the invention, described
The test strips of feline leukaemia virus Antibody preparation of color fluorescence microballoon mark delicately can quantitatively detect the cat in sample
Leukemia virus.
2. verified using the stability of the test strips of present case color fluorescence microballoon mark animal virus Antibody preparation：Group
Test strips after dress, respectively two weeks, surrounding, after eight weeks take out, take same test sample to detect, testing result CV% compared with
It is low, show that this test strips has good stability.
Claims (11)
1. a kind of preparation method of the antibody of color fluorescence microballoon mark, comprises the following steps：
(1) quantitative color fluorescence microspheres solution is added in cushioning liquid and diluted, ultrasonic 1.5min obtains color fluorescence microballoon
Suspension.
(2) it is rapid in the color fluorescence microballoon suspension into step (1) that activator is added dropwise, it is quickly placed into low on turbula shaker
After speed concussion 1min, it is fixed on turbula shaker, at room temperature quick concussion activation 30min.
(3) after activation terminates, by the color fluorescence microballoon suspension in step (2) at 26 DEG C, under 14000rpm, 12min is centrifuged
After remove supernatant, cleaned with the cushioning liquid in step (1), after being centrifuged again with the same terms, abandoning supernatant uses step
(4) cushioning liquid, which redissolves, in is resuspended.
(4) corresponding animal virus monoclonal antibody is taken, cushioning liquid is added and is diluted to 0.085mg/mL.
(5) the color fluorescence particle dispersion redissolved in step (3) is added in the antibody diluent in step (4), at once
1.5min, subsequent ultrasonic disperse 1.5min are shaken on vortex oscillator, is then attached on turbula shaker and shakes at room temperature
Swing reaction 1h.
(6) after reaction terminates, solution ultrasound 1min in step (5), at 26 DEG C, under 14000rpm, is removed after centrifugation 12min
Clear liquid, adds confining liquid, is equally then attached on turbula shaker concussion reaction 1h at room temperature.
(7) confining liquid cleaning will be added after solution centrifugal in step (6) again, at 26 DEG C, under 14000rpm, after centrifugation 12min
Supernatant is removed, the antibody of color fluorescence microballoon mark is can obtain.
2. a kind of preparation method of the antiviral antibody of color fluorescence microballoon mark according to claim 1, it is characterised in that：
The cushioning liquid of step (1) is 2- (N- morpholines) ethyl sulfonic acid cushioning liquid, and its pH is 6.8.
3. a kind of preparation method of the antibody of color fluorescence microballoon mark according to claim 1, it is characterised in that：Step (2)
Middle activator is EDC＆NHS activators.
4. a kind of preparation method of the antibody of fluorescent microsphere mark according to claim 1, it is characterised in that：Step (4)
Animal virus antibody, animal virus includes being not limited to CDV, canine parvovirus, hepatitis infectiosa canis virus, canine coronavirus, mad
Dog disease virus, feline leukaemia virus, marek's disease virus, the animal virus such as NDV.
5. a kind of usage right requires that preparation method described in 1~4 any one prepares kind of an antibody system for color fluorescence microballoon mark
Standby test strips are characterized in that, the structure of kit is as shown in Figure 1：Sample pad (2), sample pad are pasted in bottom plate (1) one end
One end closely crimping be coated with fluorescent microsphere mark animal virus antibody pad (3), pad (3) one end crimping nitre
Detection line T1 (5) and nature controlling line C (6), the other end connection adsorptive pads (7) of NC films are coated with acid cellulose NC films (4), NC films
Test paper is formed, test paper loads plastics and got stuck interior formation test card.
6. a kind of usage right requires the antibody system of color fluorescence microballoon mark prepared by preparation method described in 1~5 any one
Standby test strips, comprise the following steps：
(1) re-suspension liquid is added in the color fluorescence microballoon of antibody labeling, blown and beaten repeatedly, subsequent ultrasound 5min is until particle is completely molten
Solution.
(2) the color fluorescence microspheres solution of the antibody labeling in step (1) is sprayed at pad with film metal spraying machine is drawn, then existed
In baking oven 1h is dried at 37 DEG C.
(3) control line and detection line are wanted into coated antibody, respectively with after corresponding diluted, with draw film metal spraying machine by its
It is sprayed at respectively on nitrocellulose filter, 1h is dried in air dry oven at 37 DEG C.
(4) pad, sample pad are pasted in assembling, are allowed to fully bonding.
7. the test strips of the Antibody preparation of fluorescent microsphere mark according to claim 6, it is characterised in that：In step (2)
The solubility of the antibody-solutions of color fluorescence microballoon mark is 1.35mg/mL, and coated weight is 5 μ L/cm.
8. the test strips of the Antibody preparation of color fluorescence microballoon mark according to claim 6, it is characterised in that：Step
(3) the coated antibody of the control line of nitrocellulose filter is anti-mouse lgG polyclonal antibodies in, and coated weight is 0.9 μ L/cm；Detection
The coated antibody of line is the monoclonal antibody of corresponding animal virus, and coated weight is 0.6 μ L/cm.
9. the test strips of the Antibody preparation of fluorescent microsphere mark prepared by the preparation method as described in any one of claim 1~8
Application method.
10. the antibody of fluorescent microsphere mark prepared by the preparation method as described in any one of Claims 1 to 4 is in many animals disease
The application of poison detection.
11. the test strips of the Antibody preparation of fluorescent microsphere mark prepared by the preparation method as described in any one of claim 1~8
In the application of many animals Viral diagnosis.
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN107831317A (en) * | 2017-11-01 | 2018-03-23 | 杭州微瑞科技有限公司 | Canine coronavirus antibody Quantitative detection card and application method |
| CN108931636A (en) * | 2018-07-02 | 2018-12-04 | 威海纽普生物技术有限公司 | Measure the fluorescence immune chromatography detection kit and preparation method thereof of ST2 |
| CN109085358A (en) * | 2018-07-02 | 2018-12-25 | 威海纽普生物技术有限公司 | Serum amyloid A protein assay kit and production method |
| CN109490526A (en) * | 2017-09-13 | 2019-03-19 | 南京东纳生物科技有限公司 | A kind of preparation method that antibody is orientated the fluorescent microsphere probe modified and the application in immunochromatography |
| CN109991413A (en) * | 2017-12-31 | 2019-07-09 | 江苏雷森生物科技有限公司 | A kind of feline calicivirus fluorogenic quantitative detection card and detection method |
| CN112162092A (en) * | 2020-09-30 | 2021-01-01 | 北京金沃夫生物工程科技有限公司 | Novel coronavirus detection kit |
Citations (14)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5194300A (en) * | 1987-07-15 | 1993-03-16 | Cheung Sau W | Methods of making fluorescent microspheres |
| WO2000028598A1 (en) * | 1998-11-10 | 2000-05-18 | Biocrystal Limited | Methods for identification and verification |
| WO2003093511A1 (en) * | 2002-04-30 | 2003-11-13 | Merck & Co., Inc. | Human papillomavirus multiplexed assay |
| WO2006059052A2 (en) * | 2004-12-02 | 2006-06-08 | Cis Bio International | Inositol-phosphate derivatives and method of detecting inositol-1-phosphate |
| CN101701958A (en) * | 2009-03-05 | 2010-05-05 | 中国检验检疫科学研究院 | Fluorescent test paper strip capable of simultaneously testing newcastle disease and bird flu virus and application thereof |
| CN101957377A (en) * | 2010-09-17 | 2011-01-26 | 山东省农业科学院畜牧兽医研究所 | Method for preparing fluorescent antibody for detecting avian influenza virus and solid phase immunofluorescence detection kit |
| CN103743901A (en) * | 2013-12-19 | 2014-04-23 | 河北北方学院 | Colloidal gold test strip for detecting canine parvovirus |
| CN104530459A (en) * | 2014-12-10 | 2015-04-22 | 南京邮电大学 | Preparation method of polystyrene fluorescent microsphere coupled with antibody |
| CN204514933U (en) * | 2014-10-28 | 2015-07-29 | 广州天宝颂原生物科技开发有限公司 | B2M immunochromatographiassay assay quantitative detection test paper |
| CN105067811A (en) * | 2015-07-22 | 2015-11-18 | 中国农业大学 | T-2 toxin detecting product based on fluorescent microsphere immunochromatography and preparation method thereof |
| CN205280727U (en) * | 2015-10-15 | 2016-06-01 | 厦门宝太生物科技有限公司 | Fluorescence immunity chromatography test paper |
| CN205374469U (en) * | 2016-01-21 | 2016-07-06 | 成都微瑞生物科技有限公司 | Blue ear virus antibody detection card of pig |
| CN105738621A (en) * | 2014-12-11 | 2016-07-06 | 杨挥 | Test paper strip and method for detecting parvovirus through immunofluorescence chromatography technique |
| CN106248958A (en) * | 2016-07-21 | 2016-12-21 | 上海奥普生物医药有限公司 | The fluorescence immune chromatography reagent of a kind of detection by quantitative cTnI and preparation method |
-
2017
- 2017-02-22 CN CN201710095977.XA patent/CN107132348A/en active Pending
Patent Citations (14)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US5194300A (en) * | 1987-07-15 | 1993-03-16 | Cheung Sau W | Methods of making fluorescent microspheres |
| WO2000028598A1 (en) * | 1998-11-10 | 2000-05-18 | Biocrystal Limited | Methods for identification and verification |
| WO2003093511A1 (en) * | 2002-04-30 | 2003-11-13 | Merck & Co., Inc. | Human papillomavirus multiplexed assay |
| WO2006059052A2 (en) * | 2004-12-02 | 2006-06-08 | Cis Bio International | Inositol-phosphate derivatives and method of detecting inositol-1-phosphate |
| CN101701958A (en) * | 2009-03-05 | 2010-05-05 | 中国检验检疫科学研究院 | Fluorescent test paper strip capable of simultaneously testing newcastle disease and bird flu virus and application thereof |
| CN101957377A (en) * | 2010-09-17 | 2011-01-26 | 山东省农业科学院畜牧兽医研究所 | Method for preparing fluorescent antibody for detecting avian influenza virus and solid phase immunofluorescence detection kit |
| CN103743901A (en) * | 2013-12-19 | 2014-04-23 | 河北北方学院 | Colloidal gold test strip for detecting canine parvovirus |
| CN204514933U (en) * | 2014-10-28 | 2015-07-29 | 广州天宝颂原生物科技开发有限公司 | B2M immunochromatographiassay assay quantitative detection test paper |
| CN104530459A (en) * | 2014-12-10 | 2015-04-22 | 南京邮电大学 | Preparation method of polystyrene fluorescent microsphere coupled with antibody |
| CN105738621A (en) * | 2014-12-11 | 2016-07-06 | 杨挥 | Test paper strip and method for detecting parvovirus through immunofluorescence chromatography technique |
| CN105067811A (en) * | 2015-07-22 | 2015-11-18 | 中国农业大学 | T-2 toxin detecting product based on fluorescent microsphere immunochromatography and preparation method thereof |
| CN205280727U (en) * | 2015-10-15 | 2016-06-01 | 厦门宝太生物科技有限公司 | Fluorescence immunity chromatography test paper |
| CN205374469U (en) * | 2016-01-21 | 2016-07-06 | 成都微瑞生物科技有限公司 | Blue ear virus antibody detection card of pig |
| CN106248958A (en) * | 2016-07-21 | 2016-12-21 | 上海奥普生物医药有限公司 | The fluorescence immune chromatography reagent of a kind of detection by quantitative cTnI and preparation method |
Non-Patent Citations (1)
| Title |
|---|
| 解泉源等: "大肠杆菌O157:H7荧光微球免疫层析试纸条的研制", 《食品科学》 * |
Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109490526A (en) * | 2017-09-13 | 2019-03-19 | 南京东纳生物科技有限公司 | A kind of preparation method that antibody is orientated the fluorescent microsphere probe modified and the application in immunochromatography |
| CN107831317A (en) * | 2017-11-01 | 2018-03-23 | 杭州微瑞科技有限公司 | Canine coronavirus antibody Quantitative detection card and application method |
| CN109991413A (en) * | 2017-12-31 | 2019-07-09 | 江苏雷森生物科技有限公司 | A kind of feline calicivirus fluorogenic quantitative detection card and detection method |
| CN108931636A (en) * | 2018-07-02 | 2018-12-04 | 威海纽普生物技术有限公司 | Measure the fluorescence immune chromatography detection kit and preparation method thereof of ST2 |
| CN109085358A (en) * | 2018-07-02 | 2018-12-25 | 威海纽普生物技术有限公司 | Serum amyloid A protein assay kit and production method |
| CN112162092A (en) * | 2020-09-30 | 2021-01-01 | 北京金沃夫生物工程科技有限公司 | Novel coronavirus detection kit |
| WO2022068641A1 (en) * | 2020-09-30 | 2022-04-07 | 北京金沃夫生物工程科技有限公司 | Kit for detecting novel coronavirus, and preparation method therefor and detection method therefor |
| EP4063865A4 (en) * | 2020-09-30 | 2023-07-19 | Beijing Jinwofu Bioengineering Technology Co., Ltd | Kit for detecting novel coronavirus, and preparation method therefor and detection method therefor |
| CN112162092B (en) * | 2020-09-30 | 2024-04-02 | 北京金沃夫生物工程科技有限公司 | Novel coronavirus detection kit |
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