CN103529214B - Method for detecting treponema pallidum, quantum-dot labeled immunochromatography test paper and preparation method thereof - Google Patents
Method for detecting treponema pallidum, quantum-dot labeled immunochromatography test paper and preparation method thereof Download PDFInfo
- Publication number
- CN103529214B CN103529214B CN201310485058.5A CN201310485058A CN103529214B CN 103529214 B CN103529214 B CN 103529214B CN 201310485058 A CN201310485058 A CN 201310485058A CN 103529214 B CN103529214 B CN 103529214B
- Authority
- CN
- China
- Prior art keywords
- quantum dot
- microspironema pallidum
- monoclonal antibody
- labeled
- glass fibre
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Classifications
-
- 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
- G01N33/571—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses for venereal disease, e.g. syphilis, gonorrhoea
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Immunology (AREA)
- Engineering & Computer Science (AREA)
- Hematology (AREA)
- Chemical & Material Sciences (AREA)
- Urology & Nephrology (AREA)
- Biomedical Technology (AREA)
- Molecular Biology (AREA)
- Microbiology (AREA)
- Physics & Mathematics (AREA)
- Biotechnology (AREA)
- Virology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Food Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Cell Biology (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Peptides Or Proteins (AREA)
Abstract
The invention relates to a medical immunodetection method and particularly relates to a method for using quantum-dot labeled immunochromatography test paper to detect treponema pallidum by an immunological method. The quantum-dot labeled immunochromatography test paper is characterized in that a plastic board is stuck with a glass cellulose membrane A, a glass cellulose membrane B of a quantum-dot labeled treponema pallidum IgG monoclonal antibody, a nitrocellulose membrane and water absorbing paper from bottom to top in sequence, wherein one end of the nitrocellulose membrane is provided with treponema pallidum polyclonal antibody and rabbit antimouse second antibody so as to form a detecting band T and a quality control band C; the quantum-dot labeled treponema pallidum IgG monoclonal antibody is positioned at one end of the glass cellulose membrane and corresponds to the detecting band T and the quality control band C, and the quantum-dot labeled treponema pallidum IgG monoclonal antibody is positioned at one end of a sampling point. The method has the advantages that high specificity of immunoreactions and the fluorescence characteristic of quantum dots are combined, so that the detection sensitivity is about 1000 times higher than that of the current commonly-used colloidal gold detection method.
Description
Technical field
The present invention relates to medimmune inspection method, particularly relate to use quantum dot-labeled immunochromatographyassay assay test, with the method for immunologic methods for detecting treponema pallidum.
Background technology
Microspironema pallidum (TreponemaPallidun) is the pathogen of syphilis, because it is transparent, and not easy coloring, therefore also known as spirochaeta pallida.Syphilis is a kind of pandemic venereal disease, gos up to some extent again at the Chinese incidence of disease.Microspironema pallidum only infects the mankind, point acquired syphilis and congenital syphilis.Acquired syphilis is mainly through sexual transmission; Congenital syphilis by placenta by microspironema pallidum, is passed to fetus from Cord blood circulation, can be caused fetus general infection.Conveyor screw is amount reproduction in fetus internal organ and tissue, can cause foetal death or miscarriage.
At present, what Syphilis serum test method was conventional comprises rapid plasma reagin ring-type card experiment (RPR), tolulized red unheated serum test (TRUST), microspironema pallidum gelatin aggegation experiment (TPPA), colloidal gold method etc.What RPR, TRUST checked is microspironema pallidum specificity, sensitivity and specificity all poor (false positive and false negative are up to 30%); Although colloidal gold method detects fast and convenient, easily operate, accuracy rate is lower, and sensitivity is also lower; Although and TPPA sensitivity and specificity higher, the natural helix isoantigen that uses source difficulty, expensive, complicated operation, judged result is comparatively large by Subjective Factors, compares and is difficult to realize robotization.The detection method of therefore seeking a kind of low price, easy and simple to handle, sensitivity and specificity all higher is problem in the urgent need to address.
Summary of the invention
For the weak point of above-mentioned technology, the invention provides all higher Test paper of a kind of low price, easy and simple to handle, sensitivity and specificity and the method with this detection paper microspironema pallidum.
A kind of quantum dot-labeled immunochromatographyassay assay test, is provided with plastic plate, nitrocellulose filter, glass fibre element film A, the glass fibre element film B of quantum dot-labeled microspironema pallidum monoclonal antibody, thieving paper;
Wherein, described plastic plate is pasted with successively glass fibre element film A, the glass fibre element film B of quantum dot-labeled microspironema pallidum monoclonal antibody, nitrocellulose filter, thieving paper;
Wherein, described nitrocellulose filter one end has microspironema pallidum polyclonal antibody and rabbit against murine two to resist, and forms detection zone T and quality control band C with this;
Wherein, described quantum dot-labeled microspironema pallidum monoclonal antibody is positioned at one end of glass fibre element film B, corresponding with detection zone T and quality control band C, and quantum dot-labeled microspironema pallidum monoclonal antibody is positioned at sample delivery point one end.
Preferably, described quantum dot is CdTe/ZnSe core-shell quanta dots.
Preferably, described plastic plate is viscosity PVC base plate.
Preferably, the concentration of described microspironema pallidum polyclonal antibody is 0.5g/L.
Preferably, the concentration that described rabbit against murine two is anti-is 1.0g/L.
Preferably, described detection zone T and quality control band C spacing are no less than 5mm.
The preparation method of test paper as above, comprises the steps:
(1) coupling of quantum dot and microspironema pallidum monoclonal antibody:
Get PBS damping fluid 100 ~ 200uL and the surperficial quantum dot being connected with carboxyl of 5 ~ 20uL of 0.01M;
Choose coupling reagent, coupling reagent is selected from hydroxy succinimide, 1-(3-dimethyl aminopropyl)-3 ethyl carbon diamine hydrochloride;
Add microspironema pallidum monoclonal antibody 150 ~ 200uL;
Shaking table reaction 1 ~ 4 hour;
Chromatographic column filters, centrifugal purification;
Bovine serum albumin(BSA) with 1% ~ 5% is closed;
4 DEG C of preservations;
(2) preparation of test paper:
Resist by the PBS damping fluid of 0.05 ~ 0.15M dilution microspironema pallidum polyclonal antibody and rabbit against murine two, 0.5g/L microspironema pallidum polyclonal antibody and 1.0g/L rabbit against murine two are resisted and is sprayed on nitrocellulose filter one end, form detection zone T and quality control band C, T band and C are with interval 5mm, room temperature is dried, nitrocellulose filter is put into PBS damping fluid, close stand-by for 37 DEG C, or dry rear 4 DEG C of preservations;
Quantum dot-labeled microspironema pallidum monoclonal antibody is evenly sprayed in glass fibre membrane B one end, to be with and C is with corresponding, drying at room temperature, 4 DEG C of preservations with the T formed;
Plastic plate glues note glass fibre element film A, the glass fibre element film B of quantum dot-labeled microspironema pallidum monoclonal antibody, nitrocellulose filter, thieving paper successively;
Cut into test paper with test paper cutting knife, after drying, seal preservation.
With described detection paper microspironema pallidum, comprise the steps: sample point sample one end close to microspironema pallidum monoclonal antibody on the test paper assembled, after reaction 5min, observations in uv analyzer.
Compared with prior art, the present invention has the following advantages: owing to adding core-shell quanta dots in prepared test paper, particularly have employed CdTe/ZnSe water-soluble nuclear-shell quantum dot, by water miscible quantum dot and specific by covalent coupling, the photoluminescent property of double antibodies sandwich principle incorporating quantum point is then utilized whether to detect in sample containing object.Under the irradiation of uviol lamp, by observing the photoluminescence line of detection zone and quality control band on immuno-chromatographic test paper strip, judge testing result.The fluorescent characteristic of immunoreactive high specific and quantum dot combines by the present invention, utilizes quantum dot multi-wavelength excitation, high strength fluorescent emission, and emission peak is narrow, peak shape is symmetrical, the fluorescent characteristic that stability of photoluminescence is good, establishes fast, special, easy, sensitive immunochromatography detection method.The object quantitatively detected is reached by observing fluorescence signal.The detection sensitivity height about about 1000 times of the current conventional a kind of method for quick-collaurum of detection sensitivity ratio of the method.
Accompanying drawing explanation
Fig. 1 is process chart prepared by test paper.
Embodiment
Below in conjunction with drawings and Examples, the present invention is described in further detail.
Embodiment 1: a kind of quantum dot-labeled immunochromatographyassay assay test, be provided with plastic plate, nitrocellulose filter, glass fibre element film A, the glass fibre element film B of quantum dot-labeled microspironema pallidum monoclonal antibody, thieving paper, described glass fibre element film A is the glass fibre element film not having the market of point sample is bought;
Wherein, described plastic plate is pasted with successively glass fibre element film A, the glass fibre element film B of quantum dot-labeled microspironema pallidum monoclonal antibody, nitrocellulose filter, thieving paper;
Wherein, described nitrocellulose filter one end has microspironema pallidum polyclonal antibody and rabbit against murine two to resist, and forms detection zone T and quality control band C with this;
Wherein, described quantum dot-labeled microspironema pallidum monoclonal antibody is positioned at one end of glass fibre element film B, corresponding with detection zone T and quality control band C, and quantum dot-labeled microspironema pallidum monoclonal antibody is positioned at sample delivery point one end.
Preferably, described quantum dot is CdTe/ZnSe core-shell quanta dots.
Preferably, described plastic plate is viscosity PVC base plate.
Preferably, the concentration of described microspironema pallidum polyclonal antibody is 0.5g/L.
Preferably, the concentration that described rabbit against murine two is anti-is 1.0g/L.
Preferably, described detection zone T and quality control band C spacing are no less than 5mm.
Embodiment 2: the preparation method of test paper as above, as shown in Figure 1, comprises the steps:
(1) coupling of quantum dot and microspironema pallidum monoclonal antibody:
Get PBS damping fluid 100 ~ 200uL and the surperficial quantum dot being connected with carboxyl of 5 ~ 20uL of 0.01M;
Choose coupling reagent, coupling reagent is selected from hydroxy succinimide, 1-(3-dimethyl aminopropyl)-3 ethyl carbon diamine hydrochloride;
Add microspironema pallidum monoclonal antibody 150 ~ 200uL;
Shaking table reaction 1 ~ 4 hour;
Chromatographic column filters, centrifugal purification;
Bovine serum albumin(BSA) with 1% ~ 5% is closed;
4 DEG C of preservations;
(2) preparation of test paper:
Resist by the PBS damping fluid of 0.05 ~ 0.15M dilution microspironema pallidum polyclonal antibody and rabbit against murine two, 0.5g/L microspironema pallidum polyclonal antibody and 1.0g/L rabbit against murine two are resisted and is sprayed on nitrocellulose filter one end, form detection zone T and quality control band C, T band and C are with interval 5mm, room temperature is dried, nitrocellulose filter is put into PBS damping fluid, close stand-by for 37 DEG C, or dry rear 4 DEG C of preservations;
Quantum dot-labeled microspironema pallidum monoclonal antibody is evenly sprayed in glass fibre membrane B one end, to be with T and C is with corresponding, drying at room temperature, 4 DEG C of preservations;
Plastic plate glues note glass fibre element film A, the glass fibre element film B of quantum dot-labeled microspironema pallidum monoclonal antibody, nitrocellulose filter, thieving paper successively;
Cut into test paper with test paper cutting knife, after drying, seal preservation.
Wherein, in step (1), the detection method of coupling effect is as follows:
Gel electrophoresis: adopt 0.8 Ago-Gel, voltage is 80V, after electrophoresis 15min, in uv analyzer, observes electrophoresis result.
Dot blotting: by 0.3g/L microspironema pallidum-BSA liquid point on nitrocellulose filter, 0.01mol/L phosphate buffer (the PBS containing 5%BSA is soaked in after drying, 10nmol/L, PH=7.4, containing NaCl 8.5g/L) in, remaining protein binding site on closing membrane, hatches 1h in 37 DEG C, take out after closing, PBS washs.Prepare three identical microspironema pallidum blotting membranes, dry and put into quantum dot-labeled microspironema pallidum monoclonal antibody respectively, quantum dot-BSA and quantum dot solution afterwards, in 37 DEG C of shaking tables reaction 30min, PBS washings.Observations in uv analyzer.
Embodiment 3: with described detection paper microspironema pallidum, comprises the steps: sample point sample one end close to microspironema pallidum on the test paper assembled, after reaction 5min, and observations in uv analyzer.Blank is set to the blood of PBS damping fluid and normal person.
Result judges: under C band manifests the prerequisite of red fluorescence band, and the fluorescent belt intensity of visual T band, with blank, fluorescence is more weak, and the concentration represented containing checking matter in liquid to be measured is lower.
Embodiment 4: detection paper validation verification, the treponemal liquid solution of preparation four kinds of concentration, concentration is respectively 0.5,1.0,2.0,5.0ug/L.Utilize Test paper and chemiluminescence paper box to detect sample, each Concentration Testing 6 times, measure the validity that test paper detects result.As shown in Table 1, result show the detection paper prepared with the present invention out present the positive, and the larger fluorescence intensity of solution concentration is larger.
Table one: the ELISA test strip result of various concentration microspironema pallidum
The above, be only preferred embodiment of the present invention, and professional who are familiar with this art is after understanding technological means of the present invention such as, and natural energy, according to actual needs, is changed under the teachings of the present invention.Therefore all equal changes of doing according to the present patent application the scope of the claims and modification, once should still remain within the scope of the patent.
Claims (2)
1. a quantum dot-labeled immunochromatographyassay assay test, is characterized in that, is provided with plastic plate, nitrocellulose filter, glass fibre element film A, the glass fibre element film B of quantum dot-labeled microspironema pallidum monoclonal antibody, thieving paper;
Wherein, described plastic plate is pasted with from top to bottom successively glass fibre element film A, the glass fibre element film B of quantum dot-labeled microspironema pallidum monoclonal antibody, nitrocellulose filter, thieving paper;
Wherein, described nitrocellulose filter one end has microspironema pallidum polyclonal antibody and rabbit against murine two to resist, and forms detection zone T and quality control band C with this;
Wherein, described quantum dot-labeled microspironema pallidum monoclonal antibody is positioned at one end of glass fibre element film B, corresponding with detection zone T and quality control band C, and quantum dot-labeled microspironema pallidum monoclonal antibody is positioned at sample delivery point one end;
Wherein, described quantum dot is CdTe/ZnSe core-shell quanta dots;
Described plastic plate is viscosity PVC base plate;
The concentration of described microspironema pallidum polyclonal antibody is 0.5g/L;
The concentration that described rabbit against murine two resists is 1.0g/L;
Described detection zone T and quality control band C spacing are no less than 5mm.
2. the preparation method of quantum dot-labeled immunochromatographyassay assay test as claimed in claim 1, is characterized in that, comprise the steps:
(1) coupling of quantum dot and microspironema pallidum monoclonal antibody:
Get PBS damping fluid 100 ~ 200 μ L and the surperficial quantum dot being connected with carboxyl of 5 ~ 20 μ L of 0.01M;
Choose coupling reagent;
Add microspironema pallidum monoclonal antibody 150 ~ 200 μ L;
Shaking table reaction 1 ~ 4 hour;
Chromatographic column filters, centrifugal purification;
Bovine serum albumin(BSA) with 1% ~ 5% is closed;
4 DEG C of preservations;
Wherein, the coupling reagent in step (1) is selected from hydroxy succinimide, 1-(3-dimethyl aminopropyl)-3-ethyl carbon diamine hydrochloride;
(2) preparation of test paper:
Resist by the PBS damping fluid of 0.05 ~ 0.15M dilution microspironema pallidum polyclonal antibody and rabbit against murine two, 0.5g/L microspironema pallidum polyclonal antibody and 1.0g/L rabbit against murine two are resisted and is sprayed on nitrocellulose filter one end, form detection zone T and quality control band C, T band and C are with interval 5mm, room temperature is dried, nitrocellulose filter is put into PBS damping fluid, close stand-by for 37 DEG C, or dry rear 4 DEG C of preservations;
Quantum dot-labeled microspironema pallidum monoclonal antibody is evenly sprayed in glass fibre membrane B one end, drying at room temperature, 4 DEG C of preservations;
Plastic plate glues note glass fibre element film A, the glass fibre element film B of quantum dot-labeled microspironema pallidum monoclonal antibody, nitrocellulose filter, thieving paper successively;
Cut into test paper with test paper cutting knife, after drying, seal preservation.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310485058.5A CN103529214B (en) | 2013-10-16 | 2013-10-16 | Method for detecting treponema pallidum, quantum-dot labeled immunochromatography test paper and preparation method thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310485058.5A CN103529214B (en) | 2013-10-16 | 2013-10-16 | Method for detecting treponema pallidum, quantum-dot labeled immunochromatography test paper and preparation method thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN103529214A CN103529214A (en) | 2014-01-22 |
| CN103529214B true CN103529214B (en) | 2015-07-22 |
Family
ID=49931392
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201310485058.5A Active CN103529214B (en) | 2013-10-16 | 2013-10-16 | Method for detecting treponema pallidum, quantum-dot labeled immunochromatography test paper and preparation method thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN103529214B (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108089009A (en) * | 2017-12-13 | 2018-05-29 | 福州大学 | A kind of fetal fibronectin fluorescence immune chromatography strip and preparation method thereof |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1811449A (en) * | 2006-02-23 | 2006-08-02 | 上海交通大学 | Method for detecting quantum dot mark fast immune chromatographic test paper bar |
| CN101551398A (en) * | 2008-11-26 | 2009-10-07 | 中国计量学院 | Drug residue competition-type quantum dot-labeled immunochromatography assay test-strip and observation device thereof |
| CN101825634A (en) * | 2010-05-19 | 2010-09-08 | 厦门大学附属中山医院 | Reagent strip for joint detection of syphilis specific IgM and IgG antibodies and preparation method thereof |
| CN102830229A (en) * | 2012-08-27 | 2012-12-19 | 北京新兴四寰生物技术有限公司 | Detection reagent of treponema pallidum IgM (Immunoglobulin M) antigen colloidal gold method and preparation method thereof |
| CN103163298A (en) * | 2011-12-19 | 2013-06-19 | 天津中新科炬生物制药有限公司 | Rapid detective reagent strip of treponema pallidum immunoglobulin m (IgM) antibody and preparation method thereof |
-
2013
- 2013-10-16 CN CN201310485058.5A patent/CN103529214B/en active Active
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1811449A (en) * | 2006-02-23 | 2006-08-02 | 上海交通大学 | Method for detecting quantum dot mark fast immune chromatographic test paper bar |
| CN101551398A (en) * | 2008-11-26 | 2009-10-07 | 中国计量学院 | Drug residue competition-type quantum dot-labeled immunochromatography assay test-strip and observation device thereof |
| CN101825634A (en) * | 2010-05-19 | 2010-09-08 | 厦门大学附属中山医院 | Reagent strip for joint detection of syphilis specific IgM and IgG antibodies and preparation method thereof |
| CN103163298A (en) * | 2011-12-19 | 2013-06-19 | 天津中新科炬生物制药有限公司 | Rapid detective reagent strip of treponema pallidum immunoglobulin m (IgM) antibody and preparation method thereof |
| CN102830229A (en) * | 2012-08-27 | 2012-12-19 | 北京新兴四寰生物技术有限公司 | Detection reagent of treponema pallidum IgM (Immunoglobulin M) antigen colloidal gold method and preparation method thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| CN103529214A (en) | 2014-01-22 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN102023211B (en) | Immunochromatographic test strip for full quantitative detection of C-reactive protein and preparation method thereof | |
| CN101339196A (en) | Rapid checking method for bladder cancer by quantum dot mark immunity-chromatography test paper | |
| US11371987B2 (en) | Method of amplifying detection light using light-reflecting material, in immunochromatography | |
| CN102680692A (en) | Immunochromatographic quantitative test reagent based on near-infrared fluorescent marker | |
| CN107449898B (en) | Kanamycin residue fluorescence immunochromatographic test paper and preparation method thereof | |
| CN208367017U (en) | Serum amyloid A protein immunochromatographiassay assay quantitative detection test paper | |
| KR101718485B1 (en) | Device for Detecting Colored Reaction or Fluorescence Reaction of Immunochromatography | |
| CN102680689A (en) | Kit for detecting creatine kinase isoenzyme and preparation and use methods thereof | |
| CN103529212B (en) | Method for detecting pneumonic mycoplasma, quantum dot-labeled immunochromatographic test paper and preparation method thereof | |
| CN104198702A (en) | Quantum-dot marking test strip capable of quantitatively determining multiple indexes of blood infectious diseases and preparation method and quantitative determination method thereof | |
| US20220113309A1 (en) | Detection component for blood group antigens | |
| CN103529211B (en) | Method for detecting rubella virus, quantum dot-labeled immunochromatographic test paper and preparation method thereof | |
| CN103529213B (en) | Method for preparing quantum-dot labeled immunochromatography test paper | |
| CN103529214B (en) | Method for detecting treponema pallidum, quantum-dot labeled immunochromatography test paper and preparation method thereof | |
| CN106645043A (en) | Kit and method for fast quantitatively detecting small molecule compound | |
| CN103543269B (en) | Preparation method for quantum-dot-marked immunochromatographic test paper | |
| CN103529217A (en) | Method for detecting poliovirus, quantum dot-labeled immunochromatographic test paper and preparation method thereof | |
| CN103543262B (en) | Method for detecting EB (Epstein-Barr) virus, quantum dot labelled immunochromatographic test strip and preparation method thereof | |
| CN103543268B (en) | Preparation method for quantum dot labelled immunochromatographic test strip | |
| CN205027759U (en) | Note is inspected soon to trigeminy antibody | |
| CN103529203A (en) | Method for detecting mycobacterium tuberculosis, quantum-dot labeled immunochromatography test paper and preparation method thereof | |
| CN103529216B (en) | Preparation method for quantum-dot labeled immunochromatography test paper | |
| CN108535484A (en) | A kind of time-resolved fluoroimmunoassay chromatograph test strip and preparation method quantitatively detecting fPSA in blood | |
| CN103529215A (en) | Method for detecting measles virus, quantum-dot labeled immunochromatography test paper and preparation method thereof | |
| CN103543267A (en) | Method for detecting hepatitis A virus (HAV), quantum dot labelled immunochromatographic test strip and preparation method thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant |