CN102243232A - Diagnostic reagent kit (enzyme-linked immunosorbent assay (ELISA)) for enterovirus (EV) 71-type antibody (immune globulin M (IgM)) - Google Patents
Diagnostic reagent kit (enzyme-linked immunosorbent assay (ELISA)) for enterovirus (EV) 71-type antibody (immune globulin M (IgM)) Download PDFInfo
- Publication number
- CN102243232A CN102243232A CN2010101718622A CN201010171862A CN102243232A CN 102243232 A CN102243232 A CN 102243232A CN 2010101718622 A CN2010101718622 A CN 2010101718622A CN 201010171862 A CN201010171862 A CN 201010171862A CN 102243232 A CN102243232 A CN 102243232A
- Authority
- CN
- China
- Prior art keywords
- igm
- hole
- enzyme
- behind
- reagent kit
- 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.)
- Pending
Links
Landscapes
- Peptides Or Proteins (AREA)
Abstract
The invention relates to the field of biomedicine, in particular to an enzyme-linked immunization diagnostic reagent kit for detecting an enterovirus (EV) 71-type antibody (immune globulin M (IgM)), and a preparation method and application of the diagnostic reagent kit. The probability of hand-foot-and-mouth disease and severe infection (viral encephalitis, viral cerebrospinal meningitis and pulmonary edema) caused by EV71 type is relatively higher, and case fatality rate is relatively higher and can be 10 to 25 percent. The enzyme-linked immunization diagnostic reagent kit of the EV71-IgM antibody can be used for diagnosing the infection of the EV71 type. According to related documents about the detection of the EV71-IgM, EV71 virus cultures serving as indirect enzyme-linked immuno sorbent assay (ELISA) of envelope antigens has defects in such aspects as specificity, sensitivity and stability, and due to high cultivation cost and low efficiency, a large amount of virus cannot be supplied to the market. In order to overcome the defects, the invention provides the reagent kit which is used for detecting the EV71-IgM in human blood serum, required by clinical examination, simple and convenient to operate and applicable to all medical disease control departments, and the preparation method and the application of the reagent kit. The invention has the technical scheme that: firstly, the human blood serum is added into a micro-pore plate, wherein the IgM antibody is obtained by an anti-mu chain which is pre-enveloped on the micro-pore plate, and other uncombined components are washed and removed; secondly, an enzyme labeling object is added, the EV71-IgM in the obtained IgM can be combined with the specificity of an EV71 recombinant antigen which is labeled by horse radish peroxidase (HRP), and after washing, the HRP can react with substrates which are added subsequently; and finally, the aim of detecting the EV71-IgM antibody is fulfilled.
Description
Technical field:
The present invention relates to biomedicine field, be specifically related to ELISA measuring reagent kit of a kind of detection enterovirns type 71 antibody (IgM) and its production and application.
Technical background:
It is the general name of the disease that caused by human enterovirus 71 (EV71) that the enterovirus EV 71 type infects, mainly show as hand-foot-and-mouth disease clinically, hand-foot-and-mouth disease many in the infant eruption and prevalence, part EV71 infects infant and shows as herpangina, and viral encephalitis, viral cerebrospinal meningitis, pulmonary edema, empsyxis etc. appear in the patient with severe symptoms.Cause the cause of disease of hand-foot-and-mouth disease, modal is coxsackie virus A 16 and enterovirns type 71; Infect the large percentage of the disease generation severe infection cause by enterovirns type 71, case fatality rate is also higher, and the severe cases case fatality rate can reach 10%-25%, so etiological diagnosis is significant.
Enterovirns type 71 (EV71) genome is the sub-thread positive chain RNA that contains about 7.5Kb, comprise 5 ' end noncoding region, 3 ' end noncoding region and by polyprotein that P1, P2, P3 formed, the 11 kinds of albumen of encoding altogether, wherein 4 structural proteins VP1, VP2, VP3, VP4,7 non-structural protein 2A, 2B, 2C, 3A, 3B, 3C, 3D.Wherein VP1, VP2, VP3 are exposed to the surface of virus coat, are the main antigenic variation position of EV71; The inboard that VP4 is embedded in virus coat combines with the RNA of inside, and its function class is like the anchor of virus coat, in order to the structure of fixed virus albumen.Non-structural protein 2A is division initial stage pioneer's protein, also with to stop synthetic itself desirable proteins of host cell relevant; 3C then participates in most protein cleavage reaction; 3D is a RNA polymerase, leading transcribing and replication reaction when viral RNA duplicates; Four kinds of albumen such as 2B, 2C, 3A, 3B are then relevant with duplicating of viral RNA.Because in and epitope focus mostly in VP1, so VP1 be EV71 virus mainly in and factor of determination, so VP1 albumen is considered to be best suited for setting up the antigen of ELISA method.
The clinical diagnosis of enterovirus EV 71 is main according to three aspects: the one, and clinical manifestation, the 2nd, serological evidence, the 3rd, aetology evidence.Because it is many that the enterovirus EV 71 type infects clinical manifestation, so the serology of enterovirus EV 71 detects and the aetology detection is present topmost foundation.EV71 infects and produces the semelincident immunization protection, can detect IgM and IgG antibody after the infection.IgM reagent is the important indicator that is suitable for clinical recent infection diagnosis most.EV71-IgM antibody ELISA immunity detection reagent is the serology detection method that a kind of science, convenience, economic EV-71 infect.
The existing pertinent literature report of the detection of enterovirns type 71 antibody (IgM), basically be that viral cultures with enterovirns type 71 is the indirect ELISA of antigen as envelope antigen, kit specificity, sensitivity and stable aspect remain many weak points; And because Virus culture cost height, efficient is low, in a large number supply the market.
Summary of the invention:
The technical problem to be solved in the present invention is to overcome above-mentioned deficiency, provide a kind of clinical examination be badly in need of, easy and simple to handle, be fit to ELISA measuring reagent kit of enterovirns type 71 antibody (IgM) in detected human serum that each medical disease control department uses or the blood plasma and its production and application.
The invention provides the ELISA measuring reagent kit of a kind of enterovirns type 71 antibody (IgM), its component comprises: anti-people-IgM (μ chain) monoclonal antibody is wrapped the microwell plate of quilt, recombination enterovirus 71 type antigen, positive control, negative control, sample diluent, concentrated cleaning solution (20 *), substrate solution A, substrate solution B and the stop buffer of horseradish peroxidase-labeled in advance.
Technical scheme of the present invention is: at first serum sample is joined in the microwell plate, IgM antibody in the sample is coated on anti-people-IgM (μ chain) monoclonal antibody absorption (catching) on the microwell plate in advance, unconjugated other compositions (comprising special IgG antibody) will be washed and remove, second step, add enterovirns type 71 antigen enzyme labeling thing, EV71-IgM among the captive IgM can with specific combination of EV71 recombinant antigen of wax root peroxidase labelling, other bonds not of flush away, horseradish peroxidase can and the substrate reactions of follow-up adding.Finally reach the purpose that detects EV71-IgM antibody.
Another object of the present invention has provided the preparation method of the ELISA measuring reagent kit of a kind of enterovirns type 71 antibody (IgM), and this method comprises the following steps:
1. anti-people-IgM (μ chain) monoclonal antibody is wrapped in advance by the preparation of microwell plate
1.1 bag quilt
0.05mol/L (0.05 mM/liter), pH9.6 sodium carbonate-sodium bicarbonate buffer system adds an amount of anti-people-IgM (u chain) monoclonal antibody, join in the microwell plate by every hole 100ul (microlitre) behind the mixing, placed ambient temperature overnight 2 hours for 37 ℃, get rid of liquid in the hole then, pat dry;
1.2 washing
0.01mol/L PBS-T (0.01 mM/rise dibastic sodium phosphate-sodium dihydrogen phosphate buffer system, 0.05% Tween-20), pH7.2, the amount that adds 150ul by every hole joins in the microwell plate, gets rid of then, and repeated washing once pats dry.
1.3 sealing
0.01mol/L PBS, 10% calf serum, 0.25% casein, 0.1% merthiolate, pH7.2, the amount that adds 110ul by every hole joins in the microwell plate, places 2 hours for 37 ℃, gets rid of liquid in the hole then, pats dry;
1.4 it is dry
Microwell plate after the sealing gets rid of after liquid pats dry in the hole, is positioned over hothouse (relative humidity is less than 45%) and spends the night, and the aluminium foil bag of putting into drying agent then seals, and is stored in 2～8 ℃ of environment.
2. the preparation of enzyme-labelled antigen working fluid
0.01mol/L PBS, 10% calf serum, 0.25% casein, 1/1000Proclin300, the recombination enterovirus 71 type VP1 antigen of an amount of horseradish peroxidase-labeled of adding is stored in 2～8 ℃ of environment among the buffer of pH 7.2.
3. the preparation of positive control
Get A
450The positive many parts of human serum mixings of the anti-EV71-IgM of value 〉=2.0 with 56 ℃ of water-bath heating 120 minutes, are used 0.01mol/L PBS, 20%BSA, and the buffer of pH 7.2 adjusts to A450 value＞1.0, adds 5/10000ths merthiolate, is stored in 2～8 ℃ of environment.
4. the preparation of negative control
Get negative many parts of human serum mixings of anti-EV71-IgM of A450 value＜0.1,, behind the natural cooling, add 5/10000ths merthiolate, be stored in 2～8 ℃ of environment with 56 ℃ of water-bath heating 120 minutes.
5. substrate solution A
Na
2HPO
4.12H
2O 18g
Citric acid .H2O 5g
Sodium acetate 9.25g
Acetate 1.2ml
Na
2O
2 5g
Deionized water 1000ml, pH5.0
6. substrate solution B
Earlier with 5mlDMSO (dimethyl sulfoxide) dissolving 0.25g TMB (3,3,5, the 5-tetramethyl benzidine); Get the 1000ml deionized water and add about 1ml concentrated hydrochloric acid, pH2.5 behind the mixing joins mixing in the hydrochloric acid solution with TMB solution.
7. stop buffer
Concentrated sulphuric acid 100ml
Deionized water 800ml
8.20X concentrated cleaning solution
Na
2HPO
4.12H
2O 58g
NaH
2PO
4 8g
NaCL 160g
Tween-20 10.0ml
Deionized water 1000ml adjusts pH7.2
Another purpose of the present invention has provided enterovirns type 71 antibody (IgM) assay method in a kind of human serum, and kit of the present invention can follow these steps to operation in use:
1. take out kit and put equilibrium at room temperature 30 minutes.20 * cleansing solution is done 20 times of dilutions with distilled water.
2. add sample: take out bag by plate, perform mark, stay 1 hole blank, all the other adding negative control 3 hole positive control 50ul/ holes, 2 hole and sample 50ul/ to be measured hole are in corresponding reaction plate hole, and blank does not add.Behind reaction plate light shaking mixing,, put in 37 ℃ of incubators or the water-bath and reacted 30 minutes with shrouding film shrouding.
3. wash plate: behind the incubation, the shrouding film is taken off, blotted liquid in the hole, wash plate 5 times, soaked for 30 seconds at every turn with cleansing solution.
4. enzyme-added mark working fluid: except the every hole adding 50ul enzyme mark working fluid, blank well.
5. incubation: behind shrouding film shrouding, put in 37 ℃ of incubators or the water-bath reaction 30 minutes.
6. wash plate: behind the incubation, the shrouding film is taken off, blotted liquid in the hole, wash plate 5 times, soaked for 30 seconds at every turn with cleansing solution.
7. colour developing: every hole adds substrate A, each 50ul of B liquid, and the concussion mixing behind shrouding film shrouding, is put in 37 ℃ of incubators or the water-bath and reacted 15 minutes
8. stop: every hole adds stop buffer 50ul, light shaking mixing.
9. measure: set the microplate reader ripple and be longer than 450nm (suggestion dual wavelength 450/630nm detects), measure each hole A450 value.Attention: reading in cessation reaction 30 minutes.When using single wavelength, the calibration blank well is set the microplate reader ripple and is longer than 450nm, measures each hole A450 value, when using dual wavelength 450/630nm to detect, can not establish blank well, directly measures each hole A value.
The present invention has following characteristics:
1. adopt and catch two-step approach and detect, serum has been simplified operating process without secondary dilution, has avoided the multiple disturbing factor in the indirect method, is that sensitivity and the specificity that detects is improved.
2. method of operating is simple, and is practical, is fit to medical institutions at different levels and Disease Control and Prevention Center and uses.
Embodiment:
Example 1.Anti-people-IgM (u chain) monoclonal antibody is wrapped in advance by the preparation of microwell plate
1.1 bag quilt
Na
2CO
3 1.6g
NaHCO
3 2.9g
Deionized water 1000ml,
Adjust PH9.6, add an amount of anti-people-IgM (μ chain) monoclonal antibody, join in the microwell plate by every hole 100ul (microlitre) behind the mixing, placed 2 hours for 37 ℃, ambient temperature overnight is got rid of liquid in the hole then, pats dry;
1.2 washing
Na
2HPO
4.12H
2O 3.72g
NaH
2PO
4 0.4g
NaCL 6.8g
Tween-20 0.5ml
Deionized water 1000ml,
Adjust pH7.2 and join in the microwell plate by the amount that every hole adds 150ul, get rid of then, repeated washing once pats dry.
1.3 sealing
Na
2HPO
4.12H
2O 3.72g
NaH
2PO
4 0.4g
NaCL 6.8g
Cow's serum 100.0ml
Deionized water 1000ml,
Merthiolate 1g
Adjust pH7.2, the amount that adds 110ul by every hole joins in the microwell plate, places 2 hours for 37 ℃, gets rid of liquid in the hole then, pats dry;
1.4 it is dry
Microwell plate after the sealing gets rid of after liquid pats dry in the hole, is positioned over hothouse (relative humidity is less than 45%) and spends the night, and the aluminium foil bag of putting into drying agent then seals, and is stored in 2～8 ℃ of environment.
The preparation of example 2. enzyme-labelled antigen working fluids
Na
2HPO
4.12H
2O 3.72g
NaH
2PO
4 0.4g
NaCL 6.8g
Cow's serum 100.0ml
Casein 2.5g
Calf serum 100ml
Proclin300 1ml
Deionized water 900ml,
Adjust pH 7.2, add the recombination enterovirus 71 type VP1 antigen of an amount of horseradish peroxidase-labeled, be stored in 2～8 ℃ of environment.
The preparation of example 3. positive controls
Get A
450The positive many parts of human serum mixings of the anti-EV71-IgM of value 〉=2.0 are with 56 ℃ of water-bath heating 120 minutes, behind the natural cooling, use 0.01mol/L PBS, 20%BSA, the buffer of pH 7.2 adjusts to A450 value＞1.0, the merthiolate of adding 5/10000ths is stored in 2～8 ℃ of environment.
The preparation of example 4. negative controls
Get negative many parts of human serum mixings of anti-EV71-IgM of A450 value＜0.1,, behind the natural cooling, add 5/10000ths merthiolate, be stored in 2～8 ℃ of environment with 56 ℃ of water-bath heating 120 minutes.
Example 5. substrate solution A
Na
2HPO
4.12H
2O 18g
Citric acid .H2O 5g
Sodium acetate 9.25g
Acetate 1.2ml
Na
2O
2 5g
Deionized water 1000ml, pH5.0
Example 6. substrate solution B
Earlier with 5mlDMSO (dimethyl sulfoxide) dissolving 0.25g TMB (3,3,5, the 5-tetramethyl benzidine); Get the 1000ml deionized water and add about 1ml concentrated hydrochloric acid, pH2.5 behind the mixing joins mixing in the hydrochloric acid solution with TMB solution.
Example 7. stop buffers
Dense H
2SO
4100ml
Deionized water 800ml
Example 8.20X concentrated cleaning solution
Na
2HPO
4.12H
2O 58g
NaH
2PO
4 4g
NaCL 160g
Tween-20 10.0ml
Deionized water 1000ml adjusts pH7.2
Example 9.
9.1 with the enterovirus EV 71 reverse-transcription polymerase chain reaction kit of kit of the present invention and Ministry of Public Health medical bioengineering Technical Research Center and Da development and production in Ningbo Municipal Center for Disease Control ﹠ Prevention and clinical serum 650 examples of the parallel detection in disease prevention and control center, Hebei province (wherein healthy personnel's serum 450 examples, EV71 acute stage positive serum 192 examples, A16 positive serum 18 examples), concrete outcome is as follows:
The EV71-IgM reagent that table 1 Bell Co. produces and the comparison of PCR reagent
9.2 with kit of the present invention and virosis prevention and control clinical serum 400 examples of the parallel detection of enterovirus EV 71 neutralization test reagent (wherein healthy personnel's serum 50 examples of national measles laboratory development, EV71 acute stage positive serum 230 examples, A16 positive serum 20 examples)
The EV71-IgM reagent that table 2 Bell Co. produces and the comparison of neutralization reagent
Above result shows that the present invention is used for measuring human serum enterovirns type 71 antibody (IgM) and has good sensitivity and specificity, is applicable to the aetology laboratory auxiliary diagnosis of hand-foot-and-mouth disease, and easy and simple to handle, is fit to medical treatment at different levels and disease control department and uses.
Claims (3)
1. the ELISA measuring reagent kit of an enterovirns type 71 antibody (IgM) is characterized in that this kit comprises that anti-people-IgM (μ chain) monoclonal antibody wraps the microwell plate of quilt, recombination enterovirus 71 type antigen, positive control serum, negative control sera, concentrated cleaning solution (20 *), substrate solution A, substrate solution B and the stop buffer of horseradish peroxidase-labeled in advance.
2. the preparation method of the ELISA measuring reagent kit of a kind of enterovirns type 71 antibody according to claim 1 (IgM) is characterized in that this method comprises the following steps:
2.1. anti-people-IgM (μ chain) monoclonal antibody is wrapped in advance by the preparation of microwell plate
2.1.1 bag quilt
0.05mol/L (0.05 mM/liter), pH9.6 sodium carbonate-sodium bicarbonate buffer system adds an amount of anti-people-IgM (μ chain) monoclonal antibody, join in the microwell plate by every hole 100ul (microlitre) behind the mixing, placed ambient temperature overnight 2 hours for 37 ℃, get rid of liquid in the hole then, pat dry;
2.1.2 washing
0.01mol/L PBS-T (0.01 mM/rise dibastic sodium phosphate-sodium dihydrogen phosphate buffer system, 0.05% Tween-20), pH7.2, the amount that adds 150ul by every hole joins in the microwell plate, gets rid of then, and repeated washing once pats dry.
2.1.3 sealing
0.01mol/L PBS, 10% calf serum, 0.25% casein, 0.1% merthiolate, pH7.2, the amount that adds 110ul by every hole joins in the microwell plate, places 2 hours for 37 ℃, gets rid of liquid in the hole then, pats dry;
2.1.4 it is dry
Microwell plate after the sealing gets rid of after liquid pats dry in the hole, is positioned over hothouse (relative humidity is less than 45%) and spends the night, and the aluminium foil bag of putting into drying agent then seals, and is stored in 2～8 ℃ of environment.
2.2. the preparation of enzyme-labelled antigen working fluid
0.01mol/L PBS, 10% calf serum, 0.25% casein, 1/1000 Proclin300, the recombination enterovirus 71 type VP1 antigen of an amount of horseradish peroxidase-labeled of adding is stored in 2～8 ℃ of environment among the buffer of pH 7.2.
2.3. the preparation of positive control
Get A
450The positive many parts of human serum mixings of the anti-EV71-IgM of value 〉=2.0 with 56 ℃ of water-bath heating 120 minutes, are used 0.01mol/L PBS, 20%BSA, and the buffer of pH 7.2 adjusts to A450 value＞1.0, adds 5/10000ths merthiolate, is stored in 2～8 ℃ of environment.
2.4. the preparation of negative control
Get negative many parts of human serum mixings of anti-EV71-IgM of A450 value＜0.1,, behind the natural cooling, add 5/10000ths merthiolate, be stored in 2～8 ℃ of environment with 56 ℃ of water-bath heating 120 minutes.
2.5. substrate solution A
Na
2HPO
4.12H
2O 18g
Citric acid .H2O 5g
Sodium acetate 9.25g
Acetate 1.2ml
Na
2O
2 5g
Deionized water 1000ml, pH5.0
2.6. substrate solution B
Earlier with 5mlDMSO (dimethyl sulfoxide) dissolving 0.25g TMB (3,3,5, the 5-tetramethyl benzidine); Get the 1000ml deionized water and add about 1ml concentrated hydrochloric acid, pH2.5 behind the mixing joins mixing in the hydrochloric acid solution with TMB solution.
2.7. stop buffer
Concentrated sulphuric acid 100ml
Deionized water 800ml
2.8.20X concentrated cleaning solution
Na
2HPO
4.12H
2O 58g
NaH
2PO
4 8g
NaCL 160g
Tween-20 10.0ml
Deionized water 1000ml adjusts pH7.2
3. enterovirns type 71 antibody (IgM) assay method in the human serum is characterized in that this method comprises the following steps operation:
Put equilibrium at room temperature 30 minutes 3.1. take out kit.20 * cleansing solution is done 20 times of dilutions with distilled water.
3.2. add sample: take out bag by plate, perform mark, stay 1 hole blank, all the other adding negative control 3 hole positive control 50ul/ holes, 2 hole and sample 50ul/ to be measured hole are in corresponding reaction plate hole, and blank does not add.Behind reaction plate light shaking mixing,, put in 37 ℃ of incubators or the water-bath and reacted 30 minutes with shrouding film shrouding.
3.3. wash plate: behind the incubation, the shrouding film is taken off, blotted liquid in the hole, wash plate 5 times, soaked for 30 seconds at every turn with cleansing solution.
3.4. enzyme-added mark working fluid: except the every hole adding 50ul enzyme mark working fluid, blank well.
3.5. incubation: behind shrouding film shrouding, put in 37 ℃ of incubators or the water-bath and reacted 30 minutes.
3.6. wash plate: behind the incubation, the shrouding film is taken off, blotted liquid in the hole, wash plate 5 times, soaked for 30 seconds at every turn with cleansing solution.
3.7. colour developing: every hole adds substrate A, each 50ul of B liquid, and the concussion mixing behind shrouding film shrouding, is put in 37 ℃ of incubators or the water-bath and reacted 15 minutes
3.8. stop: every hole adds stop buffer 50ul, light shaking mixing.
3.9. measure: set the microplate reader ripple and be longer than 450nm (suggestion dual wavelength 450/630nm detects), measure each hole A450 value.Attention: reading in cessation reaction 30 minutes.When using single wavelength, the calibration blank well is set the microplate reader ripple and is longer than 450nm, measures each hole A450 value, when using dual wavelength 450/630nm to detect, can not establish blank well, directly measures each hole A value.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2010101718622A CN102243232A (en) | 2010-05-14 | 2010-05-14 | Diagnostic reagent kit (enzyme-linked immunosorbent assay (ELISA)) for enterovirus (EV) 71-type antibody (immune globulin M (IgM)) |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2010101718622A CN102243232A (en) | 2010-05-14 | 2010-05-14 | Diagnostic reagent kit (enzyme-linked immunosorbent assay (ELISA)) for enterovirus (EV) 71-type antibody (immune globulin M (IgM)) |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN102243232A true CN102243232A (en) | 2011-11-16 |
Family
ID=44961397
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2010101718622A Pending CN102243232A (en) | 2010-05-14 | 2010-05-14 | Diagnostic reagent kit (enzyme-linked immunosorbent assay (ELISA)) for enterovirus (EV) 71-type antibody (immune globulin M (IgM)) |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN102243232A (en) |
Cited By (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102590505A (en) * | 2012-02-13 | 2012-07-18 | 中国疾病预防控制中心病毒病预防控制所 | Reagent kit for detecting immunoglobulin m (IgM) AlphaLISA for resisting enterovirus71 capsid protein1 |
| CN102680694A (en) * | 2012-05-24 | 2012-09-19 | 蓝十字生物药业（北京）有限公司 | Colloidal gold test strip for rapid detection of Human enterovirus 71 (EV71) IgM (immunoglobulin m) |
| CN102944677A (en) * | 2012-11-26 | 2013-02-27 | 深圳市伯劳特生物制品有限公司 | Type detection kit and preparation method thereof for hand-foot -mouth diseases (HFMD) |
| CN103267846A (en) * | 2013-06-09 | 2013-08-28 | 湖南农业大学 | ELISA (Enzyme Linked Immunosorbent Assay) kit for detecting porcine circovirus 2 (PCV2) IgM antibody |
| CN108169484A (en) * | 2017-12-20 | 2018-06-15 | 广州瑞辉生物科技股份有限公司 | EV71 virus antigen polypeptides and its IgM antibody detection kit |
| CN109270054A (en) * | 2018-04-13 | 2019-01-25 | 宁夏医科大学 | A kind of azanol tri-chlorination iron processes measurement human serum cholinesterase method |
| CN109633167A (en) * | 2018-12-20 | 2019-04-16 | 北京贝尔生物工程股份有限公司 | A kind of human serum amyloid A assay kit of highly sensitive, wide detection range |
| CN110702897A (en) * | 2019-09-20 | 2020-01-17 | 北京贝尔生物工程股份有限公司 | Stable and efficient HRP enzymatic substrate solution and preparation method and application thereof |
| CN111458498A (en) * | 2019-01-19 | 2020-07-28 | 艾维可生物科技有限公司 | Hand-foot-mouth EV71 antigen detection kit |
| CN113711038A (en) * | 2019-01-31 | 2021-11-26 | 积水医疗株式会社 | Immunoassay method for free AIM in biological sample and method for detecting NASH in subject |
| CN113728233A (en) * | 2019-01-31 | 2021-11-30 | 积水医疗株式会社 | Immunoassay method and assay kit for free AIM in biological samples |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101509003A (en) * | 2009-03-23 | 2009-08-19 | 深圳市第三人民医院 | Sequence of enterovirns type71 genome and uses thereof |
| CN101598733A (en) * | 2009-04-16 | 2009-12-09 | 北京科兴生物制品有限公司 | EV71 virus neutralizing epitope antigen detecting agent box or reagent and preparation method thereof |
| CN101609097A (en) * | 2009-04-29 | 2009-12-23 | 唐山怡安生物工程有限公司 | The competitive enzyme-linked immune detection method of EV71 neutralizing antibody, kit or reagent and preparation method thereof |
| CN101644709A (en) * | 2009-05-26 | 2010-02-10 | 厦门大学 | Method for rapidly detecting neutralizing antibody of virus and kit therefor |
-
2010
- 2010-05-14 CN CN2010101718622A patent/CN102243232A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101509003A (en) * | 2009-03-23 | 2009-08-19 | 深圳市第三人民医院 | Sequence of enterovirns type71 genome and uses thereof |
| CN101598733A (en) * | 2009-04-16 | 2009-12-09 | 北京科兴生物制品有限公司 | EV71 virus neutralizing epitope antigen detecting agent box or reagent and preparation method thereof |
| CN101609097A (en) * | 2009-04-29 | 2009-12-23 | 唐山怡安生物工程有限公司 | The competitive enzyme-linked immune detection method of EV71 neutralizing antibody, kit or reagent and preparation method thereof |
| CN101644709A (en) * | 2009-05-26 | 2010-02-10 | 厦门大学 | Method for rapidly detecting neutralizing antibody of virus and kit therefor |
Non-Patent Citations (4)
| Title |
|---|
| SHENG-YUWANG: "Early and rapid detection of enterovirus 71 infection by an IgM-capture ELISA", 《JOURNAL OF VIROLOGICAL METHODS》 * |
| YOSHIO TANO: "Rapid Serological Diagnosis of Enterovirus 71 Infection by IgM ELISA", 《LABORTATORY AND EPIDEMIOLOGY COMMUNICATIONS》 * |
| 周伯平: "肠道病毒7 1型重组VP I 蛋白的表这和EV7 1型手足口病血清学诊断方法的建立", 《全国第2届中西医结合传染病学术会议暨国家中医药管理局第1届传染病协作组会议论文汇编》 * |
| 周伯平: "肠道病毒71型重组VPl蛋白的表达和EV71型手足口病血清学诊断方法的建立", 《中华实验与临床病毒学杂志》 * |
Cited By (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102590505A (en) * | 2012-02-13 | 2012-07-18 | 中国疾病预防控制中心病毒病预防控制所 | Reagent kit for detecting immunoglobulin m (IgM) AlphaLISA for resisting enterovirus71 capsid protein1 |
| CN102680694A (en) * | 2012-05-24 | 2012-09-19 | 蓝十字生物药业（北京）有限公司 | Colloidal gold test strip for rapid detection of Human enterovirus 71 (EV71) IgM (immunoglobulin m) |
| CN102944677A (en) * | 2012-11-26 | 2013-02-27 | 深圳市伯劳特生物制品有限公司 | Type detection kit and preparation method thereof for hand-foot -mouth diseases (HFMD) |
| CN103267846A (en) * | 2013-06-09 | 2013-08-28 | 湖南农业大学 | ELISA (Enzyme Linked Immunosorbent Assay) kit for detecting porcine circovirus 2 (PCV2) IgM antibody |
| CN108169484A (en) * | 2017-12-20 | 2018-06-15 | 广州瑞辉生物科技股份有限公司 | EV71 virus antigen polypeptides and its IgM antibody detection kit |
| CN109270054A (en) * | 2018-04-13 | 2019-01-25 | 宁夏医科大学 | A kind of azanol tri-chlorination iron processes measurement human serum cholinesterase method |
| CN109633167A (en) * | 2018-12-20 | 2019-04-16 | 北京贝尔生物工程股份有限公司 | A kind of human serum amyloid A assay kit of highly sensitive, wide detection range |
| CN111458498A (en) * | 2019-01-19 | 2020-07-28 | 艾维可生物科技有限公司 | Hand-foot-mouth EV71 antigen detection kit |
| CN113711038A (en) * | 2019-01-31 | 2021-11-26 | 积水医疗株式会社 | Immunoassay method for free AIM in biological sample and method for detecting NASH in subject |
| CN113728233A (en) * | 2019-01-31 | 2021-11-30 | 积水医疗株式会社 | Immunoassay method and assay kit for free AIM in biological samples |
| CN110702897A (en) * | 2019-09-20 | 2020-01-17 | 北京贝尔生物工程股份有限公司 | Stable and efficient HRP enzymatic substrate solution and preparation method and application thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN102243232A (en) | Diagnostic reagent kit (enzyme-linked immunosorbent assay (ELISA)) for enterovirus (EV) 71-type antibody (immune globulin M (IgM)) | |
| RU2730897C1 (en) | Method of using recombinant proteins sars-cov-2 as part of a test system for elisa test with determining igm, igg, iga class antibody levels in blood serum/plasma of covid-19 patients | |
| US8114408B2 (en) | Peptide fragments reacting specifically with antibodies against highly pathogenic newcastle disease virus and uses thereof | |
| Uusi-Kerttula et al. | Comparison of human saliva and synthetic histo-blood group antigens usage as ligands in norovirus-like particle binding and blocking assays | |
| ES2764481T3 (en) | Mycoplasma pneumoniae immunological detection kit and procedure | |
| Liu et al. | Comparative research on nucleocapsid and spike glycoprotein as the rapid immunodetection targets of COVID-19 and establishment of immunoassay strips | |
| JP2009530607A (en) | Competitive enzyme-linked immunosorbent assay (C-ELISA) for flavivirus-specific antibody detection | |
| Palaniyappan et al. | Diagnostics of severe acute respiratory syndrome-associated coronavirus (SARS-CoV) nucleocapsid antigen using chicken immunoglobulin Y | |
| WO2021169664A1 (en) | Antigen for 2019 novel coronavirus and detection use thereof | |
| BR102016011318A2 (en) | SEQUENCE OF NUCLEIC ACID, RECOMBINANT ANTIGEN, DIAGNOSTIC KITS, AND USES OF THE SAME | |
| CN109374887A (en) | Bovine viral diarrhea virus antigen colloidal gold detection kit and its application | |
| WO2005106483A1 (en) | Detection of west nile virus | |
| Blanchard et al. | Development of a recombinant truncated nucleocapsid protein based immunoassay for detection of antibodies against human coronavirus OC43 | |
| US20050100883A1 (en) | Peptide-based diagnostic reagents for SARS | |
| CN111398585B (en) | Immunodiagnosis kit for specifically detecting chikungunya virus NSP1 antigen | |
| Lehmann et al. | A line immunoassay utilizing recombinant nucleocapsid proteins for detection of antibodies to human coronaviruses | |
| Direksin et al. | An immunoperoxidase monolayer assay for the detection of antibodies against swine influenza virus | |
| CN112881710B (en) | Inhibition ELISA method for detecting foot-and-mouth disease virus antibody and application thereof | |
| CN111398594B (en) | Immunodiagnosis kit for specifically detecting yellow fever virus NS1 antigen | |
| Oseni et al. | Detection by ELISA of IgM antibodies to Ross River virus in serum from patients with suspected epidemic polyarthritis | |
| TW201337268A (en) | Method of detecting infection with the hepatitis C virus | |
| ES2257095T3 (en) | SPECIFICITY IMPROVED IN THE DETECTION OF ANTI-RUBEOLA IGM ANTIBODIES. | |
| ES2304965T3 (en) | TEST. | |
| CN107064502B (en) | A kind of multiple ELISA detection kit of pig virus infectious disease serum IgG antibody and its detection method | |
| BR112017012511B1 (en) | MONOCLONAL ANTIBODY OR A FRAGMENT THEREOF, IN VITRO AND/OR EX VIVO DIAGNOSTIC METHOD FOR HMPV INFECTION AND DIAGNOSTIC KIT FOR DETECTION OF HMPV |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C02 | Deemed withdrawal of patent application after publication (patent law 2001) | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20111116 |