CN203606367U - Immunofluorescence chromatography test strip for platelet antibody detection - Google Patents
Immunofluorescence chromatography test strip for platelet antibody detection Download PDFInfo
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- CN203606367U CN203606367U CN201320361672.6U CN201320361672U CN203606367U CN 203606367 U CN203606367 U CN 203606367U CN 201320361672 U CN201320361672 U CN 201320361672U CN 203606367 U CN203606367 U CN 203606367U
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Abstract
The utility model discloses an immunofluorescence chromatography test strip for platelet antibody detection. The immunofluorescence chromatography test strip comprises a reaction film, a sample pad, an absorption pad and a base plate, wherein the reaction film is positioned on the base plate; a detection band and a quality control band are fixed on the reaction film; the sample pad is partially overlapped with one side of the reaction film and is positioned on the reaction film and the base plate; the absorption pad is partially overlapped with the other side of the reaction film and is positioned on the reaction film and the base plate; a sample injection point is arranged on each of two sides of the sample pad and is used for dropping a sample and a fluorescent marker second antibody and washing; after the reaction is executed, the fluorescent marker second antibody indicates a reaction result. Due to the mode, the immunofluorescence chromatography test strip for platelet antibody detection, which is provided by the utility model, has a simple detection process, is high in efficiency, accurate, low in cost and practical, can realize semi-quantification or quantification detection and guarantee clinical safe, effective and scientific platelet transfusion and can also save the precious platelet resources.
Description
Technical field
The utility model relates to field of medical examination, particularly relates to the immunofluorescence chromatograph test strip that a kind of platelet antibody detects.
Background technology
Platelet surface has complicated blood group antigens, comprise that ABO antigen, HLA-I antigen and HPA antigen (are present in platelet membrane glycoprotein GP II b/ III a, GP I a/ II a, in GP I b/ IX), these antigens all can stimulate body produce corresponding antibodies and cause correlation immunity disease.
Be much because platelet antibody causes take decrease of platelet and hemorrhage disease as cardinal symptom clinically, correcting decrease of platelet symptom is usually again to select the direct requirement of disease treatment scheme.The most common thrombocytopenic disease has clinically: the female blood group incompatibility thrombopenia of Inefficacy of Platelets Transfusion disease, tire, primary autoimmune thrombopenia, secondary thrombocytopenia, medicine type immune thrombocytopenia, and much other kind as diseases such as leukaemia.Want accurately, high-level efficiency, diagnose and treat blood platelet correlation immunity disease in time, all must carry out in theory platelet antigen antibody test, but realistic situation is, only a few medical institutions can carry out platelet antigen antibody test project routine clinically both at home and abroad, its immediate cause lacks simple, efficient, accurate, inexpensive exactly, and practical platelet antigen antibody test immunological testing technology.
Platelet transfusion is one of main treatment means of current thrombopenia and various diseases.The experimental results shows, carries out crossmatch and carry out infusion with the blood platelet of screening compatibility before patient's platelet transfusion, transfuse blood to be efficiently greater than 70%, and the random platelet transfusion that only abo blood group is identical is efficient lower than 30%.This has not only wasted valuable blood, and incompatible blood platelet makes to generate platelet antibody in patient body, causes immune response, causes blood transfusion invalid, also aggravation with few exceptions of patient.At present in China, and overwhelming majority of countries in the world, be the most generally random platelet transfusion clinically, patient occurs not carry out blood group antibody detection and cross matching after the invalid disease of platelet transfusion.In Britain, the minority developed countries such as France and the U.S., numerous and diverse HLA and the platelet gene detection technique of test routine of application of expensive is the sizing of HPA gene complete series spectrum antigen gene to patient HLA-I gene and blood group of thrombocyte, select the blood donor of suitable blood group, afterwards again to a few patients using monoclonal antibody set platelet antigen analytical technology (The monoclonal antibody immobilization of platelet antigen assay, MAIPA), or erythrocyte immune adsorption test (solid-phase red cell adherence, SPRCA) carry out antibody test and cross matching.But the former experimental arrangement is numerous and diverse, easily undetected again, the latter is high to experiment operator technical requirement, and result is difficult to judge sometimes.
In China, do not set up so far blood platelet blood donor storehouse, therefore China is difficult to carry out blood group genotype compatibility platelet transfusion in recent years.MAIPA technology itself is also not suitable for being applied to conventional blood group antibody detection.The SPRCA reagent price of overseas enterprise is very expensive, and the quality of domestic reagent and supply are not guaranteed.MAIPA and SPRCA can only accomplish qualitative detection simultaneously, can not accomplish quantitative detection.
Utility model content
The technical matters that the utility model mainly solves is to provide the immunofluorescence chromatograph test strip that a kind of platelet antibody detects, simple, efficient, accurate, inexpensive and practical with this ELISA test strip.
For solving the problems of the technologies described above, the technical scheme that the utility model adopts is: the immunofluorescence chromatograph test strip that provides a kind of platelet antibody to detect, comprise reaction film, sample pad, absorption pad and base plate, described reaction film is positioned on described base plate, on described reaction film, be fixed with and detect band and quality control band, one side of described sample pad and described reaction film partly overlaps and is positioned on described reaction film and described base plate, the opposite side part of described absorption pad and described reaction film is overlapping and be positioned on described reaction film and described base plate, in described sample pad, comprise that first adds sampling point and second and add sampling point, described first adds sampling point is arranged on away from described sample pad and the overlapping side of described reaction film, described second adds sampling point is arranged on and approaches described sample pad and the overlapping side of described reaction film.
In preferred embodiment of the utility model, described detection band is at least one, and described detection is brought and is fixed with anti human platelet glycoprotein monoclonal antibody.
In preferred embodiment of the utility model, described quality control band is one, on described quality control band, is fixed with human IgG, and the material of described reaction film is nitrocellulose membrane, cellulose acetate film, nylon membrane or poly tetrafluoroethylene.
The beneficial effects of the utility model are: the immunofluorescence chromatograph test strip that platelet antibody of the present utility model detects, the operation of immunofluorescence chromatographic technique is fast and convenient, can realize sxemiquantitative or quantitatively detect in conjunction with detecting instrument, testing process is simple, efficiently, accurately, inexpensive and practical, be not easy undetected, not high to operating personnel's technical requirement, can assist the diagnosis of clinical blood platelet correlation immunity disease, and before transfusing blood, compatibility experiment detects, reduce the generation of Inefficacy of Platelets Transfusion disease and the hemorrhage and death causing thereof, ensure clinical safety, effectively, the platelet transfusion of science, also can save valuable blood platelet resource simultaneously.
Accompanying drawing explanation
Fig. 1 is the structural representation of immunofluorescence chromatograph test strip one preferred embodiment of the utility model platelet antibody detection;
In accompanying drawing, the mark of each parts is as follows: 1, reaction film, and 2, sample pad, 3, absorption pad, 4, base plate, 5, detect band, 6, quality control band, 7, first add sampling point, and 8, second add sampling point.
Embodiment
Below in conjunction with accompanying drawing, preferred embodiment of the present utility model is described in detail, thereby so that advantage of the present utility model and feature can be easier to be it will be appreciated by those skilled in the art that, protection domain of the present utility model is made to more explicit defining.
Embodiment mono-:
Refer to Fig. 1, the immunofluorescence chromatograph test strip that provides a kind of platelet antibody to detect, comprise reaction film 1, sample pad 2, absorption pad 3 and base plate 4, described reaction film 1 is positioned on described base plate 4, on described reaction film 1, be fixed with to detect and be with 5 and quality control band 6, described sample pad 2 partly overlaps and is positioned on described reaction film 1 and described base plate 4 with a side of described reaction film 1, the opposite side part of described absorption pad 3 and described reaction film 1 is overlapping and be positioned on described reaction film 1 and described base plate 4, in described sample pad 2, comprise that first adds sampling point 7 and second and add sampling point 8, described first adds sampling point 7 is arranged on a side overlapping with described reaction film 1 away from described sample pad 2, described second adds sampling point 8 is arranged on and approaches described sample pad 2 side overlapping with described reaction film 1.
Embodiment bis-:
Provide described in a kind of embodiment mono-test strips for the method for platelet antibody screening:
(1) preparation of test strips
1) preparation of reaction film
Choose Millipore HF 180 tape backing films as reaction film, be cut into 30 × 1.8cm size for subsequent use.Be that 7.2 phosphate buffers are 0.5mg/ml by the concentration adjustment of human IgG with the pH of 0.01M, adding volume fraction is 1% Tween-20, uses to draw a film instrument and be sprayed on NC film surface as nature controlling line, and drawing a film amount is 0.5 μ l/cm.After specificity is mixed for the monoclonal antibody of HLA I, GP II b/ III a, GP I a/ II a, GP I b/ IX, GP IV, be that 7.2 phosphate buffers are that to regulate its concentration be 0.5mg/ml to PBS with the pH of 0.01M, adding volume fraction is 1% Tween-20, uses to draw a film instrument and be sprayed on NC film surface as detection line.Every line is spaced apart 5mm.Draw after film finishes and put into immediately 37 ℃ of baking oven dried overnight, room temperature preservation is for subsequent use.
2) assembling of test strips
Choose domestic high-quality glass fibre membrane BT100 as sample pad material, be cut into 30 × 1.8cm size for subsequent use.Choose domestic high-quality thieving paper CH37K, and it is for subsequent use to be cut into 30 × 1.8cm size.On the PVC base plate that is 30 × 6cm in specification, first paste reaction film, then take the method for overlap joint to stick on base plate sample pad and thieving paper, the length of overlap joint is 0.5mm, so that the diffusion of reactant liquor.After assembling, be cut into the wide test strips of 4mm, and be packaged in plastic casing, be put in aluminium foil bag, 4 ℃ of lower seals are preserved.
3) preparation of fluorescence reaction liquid
It is 2mg/ml that anti-human IgG antibody-solutions is regulated to its concentration with phosphate buffer, and pH is 7.5 to 8.0.Get 1ml antibody-solutions, slowly add the cy5 fluorescent dye that 100 μ l concentration are 1mg/ml, at 4 ℃, slowly stir after 3h room temperature reaction 1h again, then dialyse with the phosphate buffer that pH is 7.4, remove unreacted cy5 dyestuff.The good fluorescence antibody of recovery coupling after dislysate completes, measures after coupling efficiency and antibody concentration, at 4 ℃, keeps in Dark Place.
(2) preparation of platelet antigen
1) extract blood platelet
Gather the fresh EDTA anticoagulated whole blood of 3 person-portion O type, centrifugal 10min under the condition that speed setting is 900rpm, getting upper strata platelet rich plasma is PRP.Be centrifugal 5min under 3800rpm by this platelet rich plasma at rotating speed, abandon supernatant liquor, pressure hematocele platelet is 0.5% EDTA and the pH PBS washing that is 7.2 3 times with containing mass concentration, supernatant discarded after last washing, and with the floating hematocrit blood platelet of residual liquid on tube wall.
2) prepare platelet antigen
Add 10 times of volume lysates, in described lysate, contain pH and be the TritonX-100 that 7.4 Tris-HCl damping fluid and volume fraction are 0.5%, mix latter 4 ℃ and hatch 30 min, then be that under 10000rpm condition, after centrifugal 5min, to get supernatant liquor for subsequent use at 4 ℃, rotating speed.
(3) detecting step
1) be 18-25 ℃ by test strips and fluorescence reaction liquid balance to room temperature;
2) add 2 μ l blood platelet lysates to sample pad.
3) adding 15 μ l to contain volume fraction is that the pH of 0.5% the Tween-20 0.15M PBS that is 7.2 is to sample pad.
4) repeat a step 3).
5) add 5 μ l sample to be checked to sample pad.
6) repeating step 3) three times.
7) add the anti-human IgG of 5 μ l cy5-to sample pad.
8) repeating step 3) three times.
9) test strips is placed in to fluorescence signal and reads instrument, read fluorescence signal and measure.
Embodiment tri-:
Provide described in a kind of embodiment mono-test strips for the method for platelet antibody specificity identification:
(1) preparation of test strips
1) preparation of reaction film
Choose Millipore HF 180 tape backing films as reaction film, be cut into 30 × 1.8cm size for subsequent use.Be that 7.2 phosphate buffers are 0.5mg/ml by the concentration adjustment of human IgG with the pH of 0.01M, adding volume fraction is 1% Tween-20, uses to draw a film instrument and be sprayed on NC film surface as nature controlling line, and drawing a film amount is 0.5 μ l/cm.Monoclonal antibody by specificity for HLA I, GP II b/ III a, GP I a/ II a, GP I b/ IX, GP IV, it is 0.5mg/ml that the 0.01M PBS that is 7.2 with pH respectively regulates its concentration, adding volume fraction is 1% Tween-20, use a stroke film instrument to be sprayed on NC film surface as detection line, every line is spaced apart 5mm, i.e. every fixing not homospecific monoclonal antibody in NC film surface.Draw after film finishes and put into immediately 37 ℃ of baking oven dried overnight, room temperature preservation is for subsequent use.
2) assembling of test strips
Choose domestic high-quality glass fibre membrane BT100 as sample pad material, be cut into 30 × 1.8cm size for subsequent use.Choose domestic high-quality thieving paper CH37K, and it is for subsequent use to be cut into 30 × 1.8cm size.On the PVC base plate that is 30 × 6cm in specification, first paste reaction film, then take the method for overlap joint to stick on base plate sample pad and thieving paper, the length of overlap joint is 0.5mm, so that the diffusion of reactant liquor.After assembling, be cut into the wide test strips of 4mm, and be packaged in plastic casing, be put in aluminium foil bag, 4 ℃ of lower seals are preserved.
3) preparation of fluorescence reaction liquid
It is 2mg/ml that anti-human IgG antibody-solutions is regulated to its concentration with PBS, and pH is 7.5 to 8.0.Get 1ml antibody-solutions, slowly add the cy5 fluorescent dye that 100 μ l concentration are 1mg/ml, at 4 ℃, slowly stir after 3h room temperature reaction 1h again, then dialyse with the phosphate buffer that pH is 7.4, remove unreacted cy5 dyestuff.The good fluorescence antibody of recovery coupling after dislysate completes, measures after coupling efficiency and antibody concentration, at 4 ℃, keeps in Dark Place.
(2) preparation of platelet antigen
1) extract blood platelet
Gather the fresh EDTA anticoagulated whole blood of 3 person-portion O type, centrifugal 10min under the condition that speed setting is 900rpm, getting upper strata platelet rich plasma is PRP.Be centrifugal 5min under 3800rpm by this platelet rich plasma at rotating speed, abandon supernatant liquor, pressure hematocele platelet is 0.5% EDTA and the pH PBS washing that is 7.2 3 times with containing mass concentration, supernatant discarded after last washing, and with the floating hematocrit blood platelet of residual liquid on tube wall.
2) prepare platelet antigen
Add 10 times of volume lysates, in described lysate, contain pH and be the TritonX-100 that 7.4 Tris-HCl damping fluid and volume fraction are 0.5%, mix latter 4 ℃ and hatch 30 min, then be that under 10000rpm condition, after centrifugal 5min, to get supernatant liquor for subsequent use at 4 ℃, rotating speed.
(3) detecting step
1) take out the fixing not test strips of homospecificity Monoclonal Antibody against Platelet, and be 18-25 ℃ by itself and fluorescence reaction liquid balance to room temperature;
2) add 2 μ l blood platelet lysates to sample pad.
3) adding 15 μ l to contain volume fraction is that the pH of the 0.5% Tween-20 0.15M PBS that is 7.2 is to sample pad.
4) repeat a step 3).
5) add 5 μ l sample to be checked to sample pad.
6) repeating step 3) three times.
7) add the anti-human IgG of 5 μ l cy5-to sample pad.
8) repeating step 3) three times.
9) test strips is placed in to fluorescence signal and reads instrument, read fluorescence signal and measure.
Embodiment tetra-:
Provide described in a kind of embodiment mono-test strips for the method for blood platelet crossmatch:
(1) preparation of test strips
1) preparation of reaction film
Choose Millipore HF 180 tape backing films as reaction film, be cut into 30 × 1.8cm size for subsequent use.Be that 7.2 phosphate buffers are 0.5mg/ml by the concentration adjustment of human IgG with the pH of 0.01M, adding volume fraction is 1% Tween-20, uses to draw a film instrument and be sprayed on NC film surface as nature controlling line, and drawing a film amount is 0.5 μ l/cm.After specificity is mixed for the monoclonal antibody of HLA I, GP II b/ III a, GP I a/ II a, GP I b/ IX, GP IV, be that to regulate its concentration be 0.5mg/ml to 7.2 PBS with the pH of 0.01M, adding volume fraction is 1% Tween-20, use a stroke film instrument to be sprayed on NC film surface as detection line, every line is spaced apart 5mm.Draw after film finishes and put into immediately 37 ℃ of baking oven dried overnight, room temperature preservation is for subsequent use.
2) assembling of test strips
Choose domestic high-quality glass fibre membrane BT100 as sample pad material, be cut into 30 × 1.8cm size for subsequent use.Choose domestic high-quality thieving paper CH37K, and it is for subsequent use to be cut into 30 × 1.8cm size.On the PVC base plate that is 30 × 6cm in specification, first paste reaction film, then take the method for overlap joint to stick on base plate sample pad and thieving paper, the length of overlap joint is 0.5mm, so that the diffusion of reactant liquor.After assembling, be cut into the wide test strips of 4mm, and be packaged in plastic casing, be put in aluminium foil bag, 4 ℃ of lower seals are preserved.
3) preparation of fluorescence reaction liquid
It is 2mg/ml that anti-human IgG antibody-solutions is regulated to its concentration with PBS, and pH is 7.5 to 8.0.Get 1ml antibody-solutions, slowly add the cy5 fluorescent dye that 100 μ l concentration are 1mg/ml, at 4 ℃, slowly stir after 3h room temperature reaction 1h again, then dialyse with the PBS that pH is 7.4, remove unreacted cy5 dyestuff.The good fluorescence antibody of recovery coupling after dislysate completes, measures after coupling efficiency and antibody concentration, at 4 ℃, keeps in Dark Place.
(2) detecting step
1) be 18-25 ℃ by test strips and fluorescence reaction liquid balance to room temperature;
2) preparation of blood donor's platelet suspension.Gather respectively the fresh EDTA anticoagulated whole blood of blood donor, speed setting is that 900rpm carries out centrifugal 10min, and getting upper strata platelet rich plasma is PRP.Be centrifugal 5min under 3800rpm by this platelet rich plasma at rotating speed, abandon supernatant liquor, pressure hematocele platelet is 0.5% EDTA and the pH PBS washing that is 7.2 3 times with containing mass concentration, is finally about 200 × 10 by PBS/EDTA damping fluid suspension blood platelet to concentration
9/ L.
3) get respectively the above-mentioned blood donor's platelet suspension of 50 μ l in test tube, respectively add 50 μ l clinical samples, mix latter 37 ℃ and hatch 30min.With PBS/EDTA damping fluid washing platelet 3 times, after last washing, discard supernatant liquor, and with the floating hematocrit blood platelet of residual liquid on tube wall.
4) add 10 times of volume lysates, in described lysate, contain pH and be the TritonX-100 that 7.4 Tris-HCl damping fluid and volume fraction are 0.5%, mix latter 4 ℃ and hatch 30 min, then be that under 10000rpm condition, after centrifugal 5min, to get supernatant liquor for subsequent use at 4 ℃, rotating speed.
5) add 2 μ l blood platelet lysates to sample pad.
6) adding 15 μ l to contain volume fraction is that the pH of the 0.5% Tween-20 0.15M PBS that is 7.2 is to sample pad.
7) repeating step 6) once.
8) add the anti-human IgG of 5 μ l cy5-to sample pad.
9) repeating step 6) three times.
10) test strips is placed in to fluorescence signal and reads instrument, read fluorescence signal and measure.
The foregoing is only embodiment of the present utility model; not thereby limit the scope of the claims of the present utility model; every equivalent structure or conversion of equivalent flow process that utilizes the utility model instructions and accompanying drawing content to do; or be directly or indirectly used in other relevant technical fields, be all in like manner included in scope of patent protection of the present utility model.
Claims (3)
1. the immunofluorescence chromatograph test strip that platelet antibody detects, it is characterized in that, comprise reaction film, sample pad, absorption pad and base plate, described reaction film is positioned on described base plate, on described reaction film, be fixed with and detect band and quality control band, one side of described sample pad and described reaction film partly overlaps and is positioned on described reaction film and described base plate, the opposite side part of described absorption pad and described reaction film is overlapping and be positioned on described reaction film and described base plate, in described sample pad, comprise that first adds sampling point and second and add sampling point, described first adds sampling point is arranged on away from described sample pad and the overlapping side of described reaction film, described second adds sampling point is arranged on and approaches described sample pad and the overlapping side of described reaction film.
2. test strips according to claim 1, is characterized in that, described detection band is at least one, and described detection is brought and is fixed with anti human platelet glycoprotein monoclonal antibody.
3. test strips according to claim 1, is characterized in that, described quality control band is one, on described quality control band, is fixed with human IgG, and the material of described reaction film is nitrocellulose membrane, cellulose acetate film, nylon membrane or poly tetrafluoroethylene.
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| CN201320361672.6U CN203606367U (en) | 2013-06-24 | 2013-06-24 | Immunofluorescence chromatography test strip for platelet antibody detection |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103336122A (en) * | 2013-06-24 | 2013-10-02 | 中国科学院苏州生物医学工程技术研究所 | Immunofluorescence chromatography test strip for detecting platelet antibody and detection method of platelet antibody |
| CN106872705A (en) * | 2016-12-30 | 2017-06-20 | 中国科学院苏州生物医学工程技术研究所 | A kind of platelet antibody specificity discrimination method and its kit |
-
2013
- 2013-06-24 CN CN201320361672.6U patent/CN203606367U/en not_active Expired - Lifetime
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103336122A (en) * | 2013-06-24 | 2013-10-02 | 中国科学院苏州生物医学工程技术研究所 | Immunofluorescence chromatography test strip for detecting platelet antibody and detection method of platelet antibody |
| CN103336122B (en) * | 2013-06-24 | 2016-01-20 | 中国科学院苏州生物医学工程技术研究所 | The immunofluorescence chromatograph test strip that a kind of platelet antibody detects and detection method |
| CN106872705A (en) * | 2016-12-30 | 2017-06-20 | 中国科学院苏州生物医学工程技术研究所 | A kind of platelet antibody specificity discrimination method and its kit |
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Effective date of registration: 20160726 Address after: Science and Technology City kolding road high tech Zone of Suzhou City, Jiangsu Province, No. 88 215163 Patentee after: SUZHOU GUOKE MEDICAL TECHNOLOGY DEVELOPMENT Co.,Ltd. Address before: Science and Technology City kolding road high tech Zone of Suzhou City, Jiangsu Province, No. 88 215163 Patentee before: Suzhou Institute of Biomedical Engineering and Technology Chinese Academy of Sciences |
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Effective date of registration: 20160914 Address after: 215163 No. 8 Jinfeng Road, Suzhou hi tech Development Zone, Jiangsu Patentee after: SUZHOU GUOKE SIBEIDA BIOTECHNOLOGY CO.,LTD. Address before: Science and Technology City kolding road high tech Zone of Suzhou City, Jiangsu Province, No. 88 215163 Patentee before: SUZHOU GUOKE MEDICAL TECHNOLOGY DEVELOPMENT Co.,Ltd. |
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Effective date of registration: 20230515 Address after: 215163 Suzhou 88 high tech Zone, Jiangsu science and Technology City Patentee after: Suzhou Guoke medical technology development (Group) Co.,Ltd. Address before: 215163 No.8 Jinfeng Road, Suzhou high tech Zone, Jiangsu Province Patentee before: SUZHOU GUOKE SIBEIDA BIOTECHNOLOGY CO.,LTD. |
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