CN101183106A - Device for detecting hepatitis b virus marker in mouth cavity liquid - Google Patents
Device for detecting hepatitis b virus marker in mouth cavity liquid Download PDFInfo
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- CN101183106A CN101183106A CNA2007100533379A CN200710053337A CN101183106A CN 101183106 A CN101183106 A CN 101183106A CN A2007100533379 A CNA2007100533379 A CN A2007100533379A CN 200710053337 A CN200710053337 A CN 200710053337A CN 101183106 A CN101183106 A CN 101183106A
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Abstract
The present invention provides a detecting device for the hepatitis B virus marker in oral cavity liquid, which relates to a device which can quickly detect the hepatitis B virus marker in the oral cavity liquid. The present invention consists of an external lid which is a oral cavity liquid gathering part, a shell upper layers of test paper which are overlapped together, a piece of test paper and the lower layer of the test paper. The present invention comprises the oral cavity liquid sample gathering part which can complete by directly gathering the oral cavity liquid through putting the paper into mouth or by using a gatherer, and reaction and result observation part which comprises antigen, antibody reaction and observation part. If the gathered oral cavity liquid has corresponding antigen, a composite is formed by the antigen, golden dot antibody, and over-membrane, a red line appears positive (sandwich method). The corresponding antibody can be detected in the same way. Or the antigen and the antibody can be detected by a competition method. And the detecting sensitivity of the sample is obviously increased by combining biotin and affinity element amplification system. The present invention can detect the hepatitis b virus marker in the oral cavity liquid quickly and conveniently and is provided with high sensitivity and specificity.
Description
Technical field
The present invention relates to a kind of can fast detecting in the mouth cavity liquid hepatitis b virus marker, and can detect the hepatitis B antigen of many index and the device of antibody simultaneously.
Background technology
Detect hepatitis type B virus (hereinafter to be referred as hepatitis B) mark--antigen and antibody in the mouth cavity liquid at present, it is used mostly to be the experimental study personnel, does not see the report that application is arranged on the clinical medicine as yet.The method that adopts is enzyme linked immunosorbent assay (Elisa), revises Elisa and radioimmunology (RIA).These methods need complicated instrument and equipment, and operation steps is cumbersome, needs the professional to operate, and spend expensively, and the time is long, and the result need wait a few hours to a couple of days usually, and radioactive material confrontation environment has pollution in the RIA method.
Mouth cavity liquid is unlike blood preparation, it is the low capacity secretion, mucin and other viscosity composition that high concentration is wherein arranged are so the fast immune chromatographic test paper method of the hepatitis b virus marker in the existing detection blood preparation (as gold mark method) is not suitable for being applied to the detection of mouth cavity liquid sample.Also do not see similar report and application are arranged.
Therefore, the device for fast detecting of developing hepatitis B mark in a kind of mouth cavity liquid is significant, and this is not only the needs of quick diagnosis, and for guidance and help hepatitis B patient's diagnosis and treatment, more back and prevent this infectious disease aspect to benefit.
Summary of the invention
The objective of the invention is at above-mentioned present situation, aim to provide a kind of can fast detecting in the mouth cavity liquid hepatitis b virus marker, and can detect the pick-up unit of hepatitis b virus marker in a kind of mouth cavity liquid of the hepatitis B antigen of many index and antibody simultaneously.
The implementation of the object of the invention is, the pick-up unit of hepatitis b virus marker in a kind of mouth cavity liquid, divide enclosing cover 1 and the test strips shell upper strata 4, test strips 6 and the test strips shell lower floor 5 that are superimposed together to form by the mouth cavity liquid collection unit, test strips shell upper strata 4 is supported by test strips shell upper layer bracket 9, and mouth cavity liquid sample slot 3, reaction and view window 7, handgrip part 8 are as a result arranged on it; Mouth cavity liquid collecting pad 2, antigen gold mark pad 10, antibody gold label pad 11 are arranged on the test strips 6, and antigen index line 12, antibody index line 13, Detection of antigen 1 district 16, Detection of antigen 2 districts 17, antibody test 1 district 18, antibody test 2 districts 19, antibody test 3 districts 20, antigen control line 14, antibody control line 15, antigen absorption pad 21, antibody absorption pad 22 and test strips antigen-antibody central authorities are every 23; Central authorities of test strips shell lower floor are arranged every 24 in the test strips shell lower floor 5.
Use when of the present invention, antigen gold mark pad 10 is set at gold anti-S of mark and gold mark anti-e potpourri (sandwich method), or golden mark HBsAg and gold mark HBeAg potpourri (striving method unexpectedly), antibody gold label pad 11 is gold mark SPA or anti-human IgG antibody, can also be gold mark HBsAg, HBeAg and HBcAg (dual-antigen sandwich method or strive method unexpectedly), or use the golden labeling antibody-anti-S of concentration known, anti-e and anti-c (striving method unexpectedly).Detection of antigen 1 district 16, Detection of antigen 2 districts 17 are respectively another anti-S, and anti-e is fixed on the film, when having corresponding antigens to exist in the sample, and antigen, golden labeling antibody, red lines positive (double antibody sandwich method) appear in antibody complex formation on the film.Also can strive insolubilized antibody (striving method unexpectedly) on the binding film unexpectedly with the antigen of concentration known and the antigen in the sample to be checked, then red lines disappear positive.When antibody test 1 district 18, antibody test 2 districts 19, antibody test 3 districts 20 detection antibody, use antigen (HBsAg respectively, HBeAg, HBcAg) the some film forms solid phase antigen, when gold mark pad is SPA, when anti-human IgG antibody or gold mark corresponding antigens,, red lines occur and represent the positive if measured antibody then forms compound when existing.If for striving method unexpectedly, 18,19,20 be respectively anti-s, anti-e, anti-c are fixed in and are capture antibody on the film, and gold mark pad then is gold mark antigen (HBsAg, HBeAg, HBcAg), when antibody to be measured exists, combine corresponding gold mark antigen with capture antibody competition on the film, the red lines of result disappear positive, show to have antibody to be checked.Also can adopt gold mark hbv antibody in the competition law.
Quote biotin and Avidin in the test strips 6,--Streptavidin or biotin--Avidin as biotin, when gold mark pad is the anti-e potpourri of gold anti-S of mark and gold mark (sandwich method), in advance with Streptavidin point membranous antigen detection zone 16,17, after film is done directly with the anti-S of biotinylation, anti-e lines on the Streptavidin, to detect corresponding antigens; Same principle detects antibody (as competition law), in advance with Streptavidin point membrane antibody detection zone 18,19,20, and can be with the anti-S of biotinylation after waiting film to do, anti-e, anti-c directly is put on the Streptavidin, and all the other are the same, and the result observes red lines and disappears positive.
Can adopt Streptavidin in addition is to catch line 16,17,18,19,20 on the film, and sample to be tested is reacted with the anti-S of biotinylation, anti-e and anti-c earlier, waits subsequent reactions by gold mark pad again, and the result observes the same.
Biotin and Avidin signal amplifying system also are suitable for the part of mark and the antigen on the immobilon-p, the various combination of antibody.
The present invention includes mouth cavity liquid sample collection part and reaction, the result observes part, and the former both can directly put into the oral cavity, places the gingiva tissue place to collect mouth cavity liquid (collecting OMT) mouth cavity liquid collecting pad 2; Also can gather by being equipped with independent mouth cavity liquid gatherer, collected mouth cavity liquid adds sample slot 3 as sample to be checked and detects.Above-mentioned mouth cavity liquid mostly be oral mucosa spill liquid (Oral Mucosal Transudate, OMT), it has from immunoglobulin (Ig) and cell composition abundant in the blood plasma; The latter is an antigen, and antibody response and result observe part.
The present invention utilizes immunology antigen-antibody principle, gathers the oral cavity liquid sample based on immune chromatography test paper method (golden mark method) and science, energy fast detecting hepatitis b virus marker in the mouth cavity liquid, and can detect the hepatitis B antigen and the antibody of many index simultaneously.Have high sensitivity, specificity, advantage such as quick and easy.
Description of drawings
Fig. 1 is a structural representation of the present invention
Fig. 2 is a test strips shell superstructure synoptic diagram
Fig. 3 is the test strips structural representation
Fig. 4 is a test strips shell understructure synoptic diagram
Embodiment
With reference to Fig. 1, the present invention divides enclosing cover 1 and the test strips shell upper strata 4, test strips 6 and the test strips shell lower floor 5 that are superimposed together to form by the mouth cavity liquid collection unit.
With reference to Fig. 2, test strips shell upper strata 4 is supported by test strips shell upper layer bracket 9, and mouth cavity liquid sample slot 3, reaction and view window 7, handgrip part 8 are as a result arranged on it.Sample slot 3 is above gold mark pad antigen gold mark pad 10, antibody gold label pad 11.Sample pad in this sample slot is made up of cellulose filter paper and glass fibre salt pad.
With reference to Fig. 3, mouth cavity liquid collecting pad 2, antigen gold mark pad 10, antibody gold label pad 11, antigen index line 12, antibody index line 13, Detection of antigen 1 district 16, Detection of antigen 2 districts 17, antibody test 1 district 18, antibody test 2 districts 19, antibody test 3 districts 20, antigen control line 14, antibody control line 15, antigen absorption pad 21, antibody absorption pad 22 and test strips antigen-antibody central authorities are arranged on the test strips 6 every 23.Mouth cavity liquid collecting pad 2 is a cellulose filter paper salt pad.
With reference to Fig. 4, central authorities of test strips shell lower floor are arranged in the test strips shell lower floor 5 every 24.
Use when of the present invention, antigen gold mark pad 10 is set at gold anti-S of mark and gold mark anti-e potpourri (sandwich method), or golden mark HBsAg and gold mark HBeAg potpourri (striving method unexpectedly), antibody gold label pad 11 is gold mark SPA or anti-human IgG antibody, can also be gold mark HBsAg, HBeAg and HBcAg (dual-antigen sandwich method or strive method unexpectedly), or use the golden labeling antibody-anti-S of concentration known, anti-e and anti-c (striving method unexpectedly).Detection of antigen 1 district 16, Detection of antigen 2 districts 17 are respectively another anti-S, and anti-e is fixed on the film, and when having corresponding antigens to exist in the sample, red lines positive (double antibody sandwich method) appear in antibody complex formation on antigen, golden labeling antibody, the film.Also can be with the antigen of concentration known and the insolubilized antibody (striving method unexpectedly) on the antigenic competition binding film in the sample to be checked, then red lines disappear positive.When antibody test 1 district 18, antibody test 2 districts 19, antibody test 3 districts 20 detection antibody, use antigen (HBsAg respectively, HBeAg, HBcAg) the some film forms solid phase antigen, when gold mark pad is SPA, when anti-human IgG antibody or gold mark corresponding antigens,, red lines occur and represent the positive if measured antibody then forms compound when existing.If for striving method unexpectedly, antibody test 1,2,3 districts 18,19,20 are respectively anti-s, anti-e, anti-c are fixed in and are capture antibody on the film, and gold mark pad then is gold mark antigen (HBsAg, HBeAg, HBcAg), when antibody to be measured exists, combine corresponding gold mark antigen with capture antibody competition on the film, the red lines of result disappear positive, show to have antibody to be checked.Also can adopt gold mark hbv antibody in the competition law.
Quote biotin and Avidin in the test strips 6, when--Streptavidin or biotin--Avidin as biotin, gold mark pad are the anti-e potpourri of gold anti-S of mark and gold mark (sandwich method), in advance with Streptavidin point membranous antigen detection 1,2 districts 16,17.Directly with the anti-S of biotinylation, anti-e lined on the Streptavidin after film was done, to detect corresponding antigens.Same principle detects antibody (as competition law), detects 1,2,3 districts 18,19,20 with Streptavidin point membrane antibody in advance, can be with the anti-S of biotinylation after waiting film dried, anti-e, anti-c directly is put on the Streptavidin, and all the other are the same, and the result observes red lines and disappears positive.
Can adopt Streptavidin in addition is to catch line 16,17,18,19,20 on the film, and sample to be tested is reacted with the anti-S of biotinylation, anti-e and anti-c earlier, waits subsequent reactions by gold mark pad again, and the result observes the same.
Biotin and Avidin signal amplifying system also are suitable for the part of mark and the antigen on the immobilon-p, the various combination of antibody.
Among the present invention the sample moving direction from left to right, when oral cavity liquid reached antigen index line 12, antibody index line 13, the index line color became pink by blueness, showed and successfully collected mouth cavity liquid.Antigen control line 14, antibody control line 15 are respectively anti-mouse IgG antibody (sandwich method) and anti-human IgG antibody.
After perhaps collecting mouth cavity liquid by the oral cavity liquid header, add sample slot 3 as sample to be checked and detect, the sample pad in this sample slot is made up of cellulose filter paper and glass fibre salt pad, and other detects and the result observes ditto.
With test strips 6 doublings of the present invention, other partly does corresponding change, and test strips becomes tow sides, and mouth cavity liquid collecting pad 2 becomes two, puts into the oral cavity jointly and collects mouth cavity liquid, and antigen is looked in the front, and reverse side is looked into antibody.
The present invention can directly look into individual event hepatitis B mark in the mouth cavity liquid, is respectively HbsAg, HbeAg, HbeAb, HbsAb, HbcAb, also can look into HBcAbIgG and IgM simultaneously.Both can be form arranged side by side, also can be the double-deck forms of tow sides, and IgG is looked in the front, reverse side is looked into IgM, can also extend and look into pres1 antigen, antibody and look into pres2 antigen, antibody.
The present invention includes mouth cavity liquid sample collection part and reaction, the result observes part, and the former both can directly put into the oral cavity, places the gingiva tissue place to collect mouth cavity liquid (collecting OMT) mouth cavity liquid collecting pad 2; Also can gather by being equipped with independent mouth cavity liquid gatherer, collected mouth cavity liquid mostly be oral mucosa spill liquid (OralMucosal Transudate, OMT), it has from immunoglobulin (Ig) and cell composition abundant in the blood plasma; The latter is an antigen, and antibody response and result observe part.
The present invention is described in detail in detail below for example:
With reference to Millipore ' s exploitation immune chromatography test paper Guide Book (second edition), immobilon-p adopts the said firm's product, the 40nm collaurum is available from BBI company, hepatitis B antigen (HbsAg, HbeAg, HBcAg), mouse monoclonal anti s, anti-e, anti-c is available from Fitzgerald company, another mouse monoclonal anti s, anti-e (anti-s on the immobilon-p, anti-e) available from Fourth Ring, Beijing biotech firm, bovine serum albumin(BSA) (BSA), staphylococcal protein A, biotin-N-hydroxy-Succinimideester, sheep anti-mouse igg antibody, goat anti-human igg antibody's (Sigma company product), hepatitis B mark (HbsAg to be measured, HbeAg, anti-s, anti-e, anti-c) available from Ministry of Public Health clinical examination center.
Example 1:
Adopt Roth J method to prepare the colloid gold label of mouse monoclonal anti s, anti-e, the glass fibre membrane pad uses bovine serum albumin(BSA) (BSA) to handle in advance, and preparation gold mark anti-s, anti-e mixes gold mark pad again, is placed into antigen gold mark pad 10 places.Another mouse monoclonal anti s, anti-e point film in Detection of antigen 1 district 16,17 places, Detection of antigen 2 districts, antigen control line 14, antibody control line 15 are sheep anti-mouse igg antibody and anti-human IgG antibody.Antigen index line 12, antibody index line 13, indicator are CaSO4.Gold labeled staphylococcus A proteins (40nm) is placed antibody gold label pad 11 places.On immobilon-p, put HbsAg, HBeAg and HbcAg, form antibody test 1 district 18, antibody test 2 districts 19, antibody test 3 districts 20.Mouth cavity liquid collecting pad 2 is a cellulose filter paper salt pad, is about to the plain filter paper of multi-layer fiber and is soaked in taking-up in a moment among 0.85%NaCl and the 1%BSA, and pad is done 37 ℃, 30 minutes, and adapted.Assemble each several part by Fig. 1.
Detect the hepatitis B mark: behind variable concentrations hepatitis B mark to be checked and normal person's mouth cavity liquid mixing, detect by the present invention, it is positive that the result red lines occur simultaneously at detection line and contrast lines place, the redfree lines occur or only a redness (control line) lines appear as feminine gender.The result observes and is no more than 30 minutes.The present invention successfully detects above-mentioned mark, and the least concentration HbsAg that wherein records is 3ng/ml, and HbsAb is 30mIv/ml, and HbeAg is 2NCU/ml, and HbeAb is 4NCU/ml, and HbcAb is 2NCU/ml.
Example 2:
1) prepares anti-s, anti-e and anti-c-biotin bond respectively, adopt biotin-N-hydroxySuccinimide ester combination.
2) preparing hepatitis B antigen detects: gold mark pad (anti-s, anti-e mixes) prepare with precedent 1, the trapping region film with Streptavidin 6mg/ml Tris butfer mid point film in Detection of antigen 1 district 16, Detection of antigen 2 districts 17, after waiting to do, give birth to elementization monoclonal anti s, anti-e (1.0mg/ml) and directly put on Streptavidin respectively.All the other steps are with example 1, contrast to be sheep anti-mouse igg antibody.
3) prepare hbv antibody and detect (competition law): at first prepare HBsAg, HBeAg and HBcAg and mix gold mark pad, similar above-mentioned elder generation puts film in formation antibody test 1 district 18, antibody test 2 districts 19, antibody test in 3 districts 20 with Streptavidin, prepare the anti-s of biotinylation with step 1 again, anti-e and anti-c, directly on Streptavidin, assemble each several part.Contrast is the anti-human IgG antibody of gold mark.Assemble each several part by Fig. 1.
4) test specimen: with the hepatitis B mark mixing of various variable concentrations to be measured in normal person's mouth cavity liquid.The result observes, and antigen part (Detection of antigen line) has red lines positive, looks into the red lines of antibody moiety (antibody detection line) and disappears positive.This example not only successfully detects the hepatitis B mark, and susceptibility obviously improves than example 1, and the least concentration that wherein records is that HbsAg is 1ng/ml, and HbsAb is 10mIv/ml, and HbeAg is 1NCU/ml, and HbeAb is 2NCU/ml, and HbcAb is 1NCU/ml.Compare with enzyme linked immunosorbent assay (Elisa) in addition, detect above-mentioned test specimen, the two positive findings sees the following form:
| The present invention | ELisa | |
| HBsAg | 16 | 18 |
| HBeAg | 6 | 7 |
| HBsAb | 5 | 5 |
| HBeAb | 11 | 12 |
| HBcAb | 14 | 14 |
As seen from the table, both testing results are basic identical.
Claims (5)
1. the pick-up unit of hepatitis b virus marker in the mouth cavity liquid, it is characterized in that dividing enclosing cover (1) and the test strips shell upper strata (4), test strips (6) and the test strips shell lower floor (5) that are superimposed together form by the mouth cavity liquid collection unit, test strips shell upper strata (4) is supported by test strips shell upper layer bracket (9), and mouth cavity liquid sample slot (3), reaction and view window (7), handgrip part (8) are as a result arranged on it; Mouth cavity liquid collecting pad (2), antigen gold mark pad (10), antibody gold label pad (11), antigen index line (12), antibody index line (13), Detection of antigen 1 district (16), Detection of antigen 2 districts (17), antibody test 1 district (18), antibody test 2 districts (19), antibody test 3 districts (20), antigen control line (14), antibody control line (15), antigen absorption pad (21), antibody absorption pad (22) and test strips antigen-antibody central authorities are arranged every (23) on the test strips (6); Central authorities of test strips shell lower floor are arranged every (24) in the test strips shell lower floor (5).
2. the pick-up unit of hepatitis b virus marker in a kind of mouth cavity liquid according to claim 1 is characterized in that mouth cavity liquid collecting pad (2) is a cellulose filter paper salt pad.
3. the pick-up unit of hepatitis b virus marker in a kind of mouth cavity liquid according to claim 1 is characterized in that the sample pad in the mouth cavity liquid sample slot (3) is made up of cellulose filter paper and glass fibre salt pad.
4. the pick-up unit of hepatitis b virus marker in a kind of mouth cavity liquid according to claim 1 is characterized in that quoting biotin and Avidin in the test strips (6), as biotin-Streptavidin or biotin--Avidin.
5. the pick-up unit of hepatitis b virus marker in a kind of mouth cavity liquid according to claim 1 is characterized in that test strips shell upper strata (4), test strips (6) and test strips shell lower floor (5) doubling that will be superimposed together.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA2007100533379A CN101183106A (en) | 2007-09-21 | 2007-09-21 | Device for detecting hepatitis b virus marker in mouth cavity liquid |
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| Application Number | Priority Date | Filing Date | Title |
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| CNA2007100533379A CN101183106A (en) | 2007-09-21 | 2007-09-21 | Device for detecting hepatitis b virus marker in mouth cavity liquid |
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Cited By (10)
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| CN101614739A (en) * | 2009-07-30 | 2009-12-30 | 杭州艾力康医药科技有限公司 | Hepatitis C virus antibody saliva fast detection test paper strip |
| CN101750496A (en) * | 2009-10-16 | 2010-06-23 | 北京科美东雅生物技术有限公司 | Magnetic immunochromatographic test strip for detecting hepatitis B e antibody and preparation method thereof |
| CN102313802A (en) * | 2010-07-09 | 2012-01-11 | 上海荣盛生物药业有限公司 | Component for detecting antigen or antibody and application of component |
| CN101614740B (en) * | 2009-07-30 | 2013-01-02 | 杭州艾力康医药科技有限公司 | Hepatitis A virus antigen saliva fast detection test paper strip |
| CN103175958A (en) * | 2013-02-17 | 2013-06-26 | 北京市药品检验所 | Method for quickly identifying recombinant hepatitis B vaccine |
| CN106124586A (en) * | 2016-06-20 | 2016-11-16 | 山东理工大学 | A kind of preparation method and application of the sensor of two kinds of hepatitis b virus marker HBs/HBe of detection simultaneously |
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| CN108196050A (en) * | 2018-02-02 | 2018-06-22 | 江苏维尔生物科技有限公司 | For detecting the colloid gold test paper and its preparation and application of human immune defect virus antibody and syphilis helicoid antibody in saliva |
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| CN101614739A (en) * | 2009-07-30 | 2009-12-30 | 杭州艾力康医药科技有限公司 | Hepatitis C virus antibody saliva fast detection test paper strip |
| CN101614740B (en) * | 2009-07-30 | 2013-01-02 | 杭州艾力康医药科技有限公司 | Hepatitis A virus antigen saliva fast detection test paper strip |
| CN101750496A (en) * | 2009-10-16 | 2010-06-23 | 北京科美东雅生物技术有限公司 | Magnetic immunochromatographic test strip for detecting hepatitis B e antibody and preparation method thereof |
| CN102313802A (en) * | 2010-07-09 | 2012-01-11 | 上海荣盛生物药业有限公司 | Component for detecting antigen or antibody and application of component |
| CN102313802B (en) * | 2010-07-09 | 2014-04-23 | 上海荣盛生物药业有限公司 | Component for detecting antigen or antibody and application of component |
| CN103175958A (en) * | 2013-02-17 | 2013-06-26 | 北京市药品检验所 | Method for quickly identifying recombinant hepatitis B vaccine |
| CN106124586A (en) * | 2016-06-20 | 2016-11-16 | 山东理工大学 | A kind of preparation method and application of the sensor of two kinds of hepatitis b virus marker HBs/HBe of detection simultaneously |
| CN106124586B (en) * | 2016-06-20 | 2018-08-14 | 山东理工大学 | A kind of preparation method and application of sensor that is while detecting two kinds of hepatitis b virus marker HBs/HBe |
| CN108181468A (en) * | 2018-02-02 | 2018-06-19 | 江苏维尔生物科技有限公司 | For detecting colloid gold test paper of syphilis helicoid antibody and preparation method thereof and application method in saliva |
| CN108196050A (en) * | 2018-02-02 | 2018-06-22 | 江苏维尔生物科技有限公司 | For detecting the colloid gold test paper and its preparation and application of human immune defect virus antibody and syphilis helicoid antibody in saliva |
| CN108732347A (en) * | 2018-05-23 | 2018-11-02 | 江苏维尔生物科技有限公司 | A kind of detection kit and preparation method thereof detecting HCV antibody in saliva |
| CN112114134A (en) * | 2019-06-19 | 2020-12-22 | 爱科来株式会社 | Target substance detection method, target substance detection kit, and target substance detection system |
| CN112114134B (en) * | 2019-06-19 | 2024-02-02 | 爱科来株式会社 | Target substance detection method, target substance detection kit, and target substance detection system |
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Open date: 20080521 |