CN101995464A - Method for detecting mycobacteria tuberculosis - Google Patents
Method for detecting mycobacteria tuberculosis Download PDFInfo
- Publication number
- CN101995464A CN101995464A CN2009100577732A CN200910057773A CN101995464A CN 101995464 A CN101995464 A CN 101995464A CN 2009100577732 A CN2009100577732 A CN 2009100577732A CN 200910057773 A CN200910057773 A CN 200910057773A CN 101995464 A CN101995464 A CN 101995464A
- Authority
- CN
- China
- Prior art keywords
- antibody
- tubercle bacillus
- bacteriophage
- albumen
- weight portion
- 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.)
- Granted
Links
Landscapes
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Peptides Or Proteins (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention discloses a method for detecting mycobacteria tuberculosis, which comprises the following steps of: 1) incubating and cultivating the mycobacteria tuberculosis to amplify the mycobacteria tuberculosis; 2) adding phages into the mycobacterium tuberculosis, and lysing the mycobacteria tuberculosis after the incubation; and 3) preparing a phage colloidal gold immunochromatographic assay pen by using the antibody labeled gold of the phages, and detecting the phages by using the pen. In the method of the invention, the phages of the mycobacteria tuberculosis can be quickly recognized to further reflect the number of the mycobacteria tuberculosis. Therefore, the method simplifies detection steps and greatly shortens the detection time.
Description
Technical field
The present invention relates to molecular biology, immunology and protein detection technology field.More specifically, relate to a kind of method that detects tubercle bacillus.
Background technology
Pulmonary tuberculosis is the chronic infectious disease of a kind of infecting both domestic animals and human of being caused by tubercle bacillus, is present single infective bacterial causing death's principal element.In recent years, along with increasing and the appearance of multiple drug resistance bacterial strain of popular, the multiple malignant tumor patient of acquired immune deficiency syndrome (AIDS), increased the difficulty in the control of tubercle bacillus affection disease especially.Statistics shows, there is tuberculosis germ the infected 2,000,000,000 (accounting for total population 1/3) in the whole world, infects a people p.s., annual new 8,000,000 people that infect of global tuberculosis germ, if do not take the non-proliferation measure immediately, tuberculosis will be seized more than 3,000 ten thousand people's life in 10 years from now on.China whole nation tuberculosis infection person 3.3 hundred million, tuberculosis patient 6,000,000, annual patient because of tuberculosis death is the twice of the dead summations of all kinds of infectious diseases up to 250,000, therefore China is upgraded to Category B notifiable disease with tuberculosis by the Class C infectious disease, lists the strict control category in.According to WHO statistics, the whole world has 5,000 ten thousand to 100,000,000 people to infect tuberculosis every year, and about three million peoples die from tuberculosis, wherein more than 80% in developing country.Therefore, how to diagnose and treat the key link that becomes the diffusion of control tuberculosis and propagate fast as soon as possible.
At present, phthisical diagnosis mainly is by getting sputum examination except general chest X-ray examination, comprises that acid-fast bacilli sputum smear dyeing microscopy and tubercle bacillus cultivate, and the bacteriology checking of tubercle bacillus is to make a definite diagnosis phthisical important evidence.Acid-fast bacilli plate coating checking susceptibility is low, poor specificity, and can't determine that bacterium is anyway.Tubercle bacillus is cultivated normally with the Russell medium of specimen inoculation to be checked to improvement, cultivates 20～60 days, and treats comprehensively to judge according to biological property and biochemical reaction many index behind the bacterial growth again for 37 ℃.Since the thalline complex structure of tulase, rich lipid, and hydrophobicity makes that the pulmonary bacillus speed of growth is slow, just divides once every 12～twenty four hours.So, utilize cultural method to detect the existence of tulase, often need the time in three～eight weeks just can obtain the result, length expends time in, and complex operation, can't satisfy and make a definite diagnosis phthisical needs fast, often the opportunity of delay treatment and susceptibility not high (positive rate only about 40%).
Recent years, bacteriophage rapid amplifying method is utilized the quick growth characteristics of mycobacterium smegmatis and the relative specificity of D29 bacteriophage, carries out the fast detecting of tubercle bacillus.The core content of this method is:
A. handled sample to be checked 30 minutes, the centrifugal supernatant that goes with sample preparation liquid chamber temperature;
B. increased bacterium 24 hours with 37 ℃ of enrichment liquids;
C. add 37 ℃ of incubations of D29 mycobacteriophage 1 hour, and infected the tubercle bacillus thalline;
D. do not invade the bacteriophage of tubercle bacillus with ending the liquid deactivation, room temperature effect 10 minutes is with aborting infection;
E. get the liquid to be measured of steps d gained, drip and to wrap in advance on the agar bacterium colony double dish of quilt in mycobacterium smegmatis, 37 ℃ incubation 15-20 hour;
F. according to the formation situation result of determination of plaque on the bacterium colony double dish.
The kit high specificity of this method, result are distinct, need not specific apparatus, can be used for the tubercle bacillus thalline Rapid identification of living in the various samples to be checked; But whole test needed just can finish at least in 48 hours, and operation is complicated, and sensitivity is also not high enough.
In recent years, increasingly mature along with Protocols in Molecular Biology, antigen purification technology and monoclonal antibody technique, some fast the method for inspection come out one after another, as: polymerase chain reaction method (polymerase chain reaction, PCR), enzyme linked immunosorbent assay (Enzyme linked immunosorbent assay, ELISA), spot immune gold percolation (dot immuno-gold filtration assay, DIGFA) and immunochromatographic method (immunochormatiographic test, ICT).
Polymerase chain reaction,PCR is the external enzymatic amplification technique that is mediated mycobacterium tuberculosis dna particular sequence fragment in the sample to be checked by a pair of specific Oligonucleolide primers, circulation by three different temperatures, dozens of sex change, renaturation, extension, the particular target sequence can be amplified millions of times, but and show its fast, characteristics such as high sensitivity quantitative test and theoretic high specific.But this method is operated more complicated, and higher to operating equipment and personnel's technical requirement, the laboratory easily produces pollution.
Enzyme linked immunosorbent assay, spot immune gold percolation and immunochromatographic method all are based on the detection method that sero-immunity grows up.These methods all are that the characteristic antigen with tubercle bacillus is catches, anti-human immunoglobulin(HIg) or staphylococcal protein A (Staphylococal Protein A with mark, SPA) etc. be indication mechanism, detect the against mycobacterium tuberculosis circulating antibody that exists in patient's body, indirectly patient's ill situation is diagnosed.Enzyme linked immunosorbent assay needs the instrument of specialty and the professional of high operative skill.Immunity percolation method and immunochromatographic method detect required time and also shorten dramatically all without any need for instrument, and be easy and simple to handle, visual result.But the common problem of said method is sensitivity and specificity is difficult to reach requirement, and can not distinguish the case that infection has once been cured; The product that has even can not distinguish and inject Bacille Calmette-Guerin (Bacillus Calmette-Guerin, case BCG).
Summary of the invention
The technical problem to be solved in the present invention provides a kind of method that detects tubercle bacillus, and this method can quick identification tubercle bacillus bacteriophage, and then the quantity of reflection tubercle bacillus, because simplified the detection step, has shortened detection time widely.
For solving the problems of the technologies described above, a kind of method that detects tubercle bacillus of the present invention may further comprise the steps:
1) the tubercle bacillus incubation is cultivated with the amplification tubercle bacillus;
2) in tubercle bacillus, add bacteriophage, cracking bacterium behind the incubation;
3) with phage-resistant antibody standard gold, preparation bacteriophage colloidal gold method immune chromatograph testing pen detects bacteriophage with this test pen.
The antibody of colloid gold label is for discerning the antibody of tubercle bacillus bacteriophage head albumen or tail protein in the test pen described in the step 3), and fixing antibody is the antibody of identification tubercle bacillus bacteriophage head albumen or tail protein on the p-wire.Following four kinds of modes are promptly arranged: the antibody of colloid gold label is the antibody of identification tubercle bacillus bacteriophage head albumen in the A. test pen, and fixing antibody also is the antibody of identification tubercle bacillus bacteriophage head albumen on the p-wire.B. the antibody of colloid gold label is the antibody of identification tubercle bacillus bacteriophage tail albumen in the test pen, and fixing antibody also is the antibody of identification tubercle bacillus bacteriophage tail albumen on the p-wire.C. the antibody of colloid gold label is the antibody of identification tubercle bacillus bacteriophage head albumen in the test pen, and fixing antibody is the antibody of identification tubercle bacillus bacteriophage tail albumen on the p-wire.D. the antibody of colloid gold label is the antibody of identification tubercle bacillus bacteriophage tail albumen in the test pen, and fixing antibody is the antibody of identification tubercle bacillus bacteriophage head albumen on the p-wire.
The antibody specific bond tubercle bacillus bacteriophage head albumen of described identification tubercle bacillus bacteriophage head albumen, this tubercle bacillus bacteriophage head protein fragments has the amino acid sequence shown in nucleotide sequence shown in the SEQ ID NO:1 and the SEQ ID NO.2.
The antibody specific bond tubercle bacillus bacteriophage tail albumen of described identification tubercle bacillus bacteriophage tail albumen, this tubercle bacillus bacteriophage tail protein fragments has the amino acid sequence shown in nucleotide sequence shown in the SEQ ID NO:3 and the SEQ ID NO.4.
Step 1) is specifically in two steps:
A) with 4% NaOH solution room temperature treatment sample to be checked 20 minutes, the centrifugal supernatant that goes;
B) add enrichment liquid, cultivated 18～24 hours for 37 ℃, the centrifugal supernatant that goes adds the resuspended tubercle bacillus of enrichment liquid again.
Described enrichment liquid is by Michaelis 7H9 nutrient culture media 5-7 weight portion, lime chloride 1-2 weight portion, ampicillin 25-50 weight portion, amphotericin B 25-50 weight portion, bovine serum albumin(BSA) 100-200 weight portion, glucose 10-15 weight portion, oleic acid 1-2 weight portion and hydrogen peroxidase 0.1-0.2 weight portion, all is dissolved in the pure water of 1000 weight portions being prepared from.
Compare with prior art, the present invention has following beneficial effect: can quick identification tubercle bacillus bacteriophage with method of the present invention, and then the quantity of reflection tubercle bacillus.Because neither need to kill free bacteriophage, do not need to add again shame dirt bacillus incubation yet or on agar plate incubated overnight form plaque, so not only simplified the detection step, and shortened detection time.
Embodiment
The invention will be further elaborated with contrast test by the following examples:
Embodiment 1, the preparation of the antibody of identification tubercle bacillus bacteriophage D29 head albumen
1, tubercle bacillus bacteriophage D29 head protein fragments synthetic
The nucleotide sequence of tubercle bacillus bacteriophage D29 head protein fragments is shown in SEQ ID NO:1, and the amino acid sequence of tubercle bacillus bacteriophage D29 head protein fragments is shown in SEQ ID NO:2.
The nucleic acid that method by molecular cloning will contain SEQ ID NO:1 sequence is inserted into pcDNA3 carrier (available from Invitrogen company), and order-checking is selected correct clonal expansion after identifying, extracts transformed into escherichia coli behind the DNA, expresses and the fragment of purifying D29 head albumen.
2, the preparation of the antibody of identification D29 head protein fragments
A) animal immune
D29 head protein fragments and isopyknic freund adjuvant of front purifying are mixed and made into water-in-oil emulsion, and every mouse peritoneal injection above-mentioned mixed liquor of 0.5ml (containing head protein fragments 100 μ g) is injected 6 BALB/c mouse altogether.Add the freund 's incomplete adjuvant booster immunization with doses of antigen after two weeks, detect many anti-the tiring of mice serum head albumen with indirect ELISA after 7 days, the high person's cephalic vein of tiring impacts immunity 1 time again, and every 20 μ g carries out Fusion of Cells after 3 days.
B) foundation of hybridoma cell strain
With the splenocyte suspension of immune mouse and SP2/0 myeloma cell with 5: 1 ratio under the polyglycol effect routinely method merge, with the complete 1640 nutrient culture media screening and culturing of HAT.(1 μ g/ml) marks 96 orifice plates as antigen coated enzyme with D29 head albumen, and indirect elisa method detects the Hybridoma Cell Culture supernatant, and the OD490 that is surveyed (absorption at 490nm place) carries out subcloning than the high 10 times clone of negative control, and it is frozen to increase.Through 3 limited dilution cloningizations, with a large amount of amplifications of the hybridoma cell line of secreting specificity antibody and frozen, after the cultivation of going down to posterity for a long time, with the cloning evaluation once more of identical method.
C) preparation of monoclonal antibody ascites
The BALB/c mouse intraperitoneal is injected the incomplete freund adjuvant of 0.5ml respectively, and every the interior injection of mouse peritoneal 0.5ml contains 2 * 10 after 5～7 days
6The solution of individual hybridoma, backsight mouse web portion degrees of expansion was collected ascites in 7～10 days.
D) Purification of Monoclonal Antibodies
Successively use 50% (mass volume ratio) saturated ammonium sulfate to saltout the monoclonal antibody ascites of identification D29 head albumen and the method for Protein G affinity chromatography is carried out purifying, the monoclonal anti body and function protein electrophoresis that obtains is identified purity.
Embodiment 2, the preparation of the antibody of identification tubercle bacillus bacteriophage D29 tail protein
1, tubercle bacillus bacteriophage D29 tail protein fragment synthetic
The nucleotide sequence of tubercle bacillus bacteriophage D29 tail protein fragment is shown in SEQ ID NO:3, and the amino acid sequence of tubercle bacillus bacteriophage D29 tail protein fragment is shown in SEQ ID NO:4.
The nucleic acid that method by molecular cloning will contain SEQ ID NO:1 sequence is inserted into pcDNA3 carrier (available from Invitrogen company), and order-checking is selected correct clonal expansion after identifying, extracts transformed into escherichia coli behind the DNA, expresses and purifying D29 tail protein fragment.
2, the preparation of the antibody of identification D29 tail protein fragment
A) animal immune
D29 tail protein fragment and isopyknic freund adjuvant of front purifying are mixed and made into water-in-oil emulsion, and every mouse peritoneal injection above-mentioned mixed liquor of 0.5ml (containing tail protein fragment 100 μ g) is injected 6 BALB/c mouse altogether.Add the freund 's incomplete adjuvant booster immunization with doses of antigen after two weeks, detect many anti-the tiring of mice serum tail protein with indirect ELISA after 7 days, the high person's tail vein of tiring impacts immunity 1 time again, and every 20 μ g carries out Fusion of Cells after 3 days.
B) foundation of hybridoma cell strain
With the splenocyte suspension of immune mouse and SP2/0 myeloma cell with 5: 1 ratio under the polyglycol effect routinely method merge, with the complete 1640 nutrient culture media screening and culturing of HAT.(1 μ g/ml) marks 96 orifice plates as antigen coated enzyme with the D29 tail protein, and indirect elisa method detects the Hybridoma Cell Culture supernatant, and the OD490 that is surveyed (absorption at 490nm place) carries out subcloning than the high 10 times clone of negative control, and it is frozen to increase.Through 3 limited dilution cloningizations, with a large amount of amplifications of the hybridoma cell line of secreting specificity antibody and frozen, after the cultivation of going down to posterity for a long time, with the cloning evaluation once more of identical method.
C) preparation of monoclonal antibody ascites
The BALB/c mouse intraperitoneal is injected the incomplete freund adjuvant of 0.5ml respectively, and every the interior injection of mouse peritoneal 0.5ml contains 2 * 10 after 5～7 days
6The solution of individual hybridoma, backsight mouse web portion degrees of expansion was collected ascites in 7～10 days.
D) Purification of Monoclonal Antibodies
Successively use 50% (mass volume ratio) saturated ammonium sulfate to saltout the monoclonal antibody ascites of identification D29 tail protein and the method for Protein G affinity chromatography is carried out purifying, the monoclonal anti body and function protein electrophoresis that obtains is identified purity.
Embodiment 3, amplification and the detection of D29 bacteriophage in shame dirt bacillus
1, (Au probe obtains with the antibody (embodiment 1 makes) of the anti-D29 head of colloid gold label albumen colloidal gold method immune chromatograph testing pen, and p-wire antibody is also used this antibody; Perhaps Au probe obtains with the antibody (embodiment 2 makes) of the anti-D29 tail protein of colloid gold label, p-wire antibody is also used this antibody) be 200810043709.4 with application number, the applying date is on August 14th, 2008, the method preparation of denomination of invention for being put down in writing in the Chinese invention patent application specification of " method of a kind of fast detecting tubercle bacillus and kit ".
2, adding 0.1ml concentration in well respectively is 10
9Individual/ml, 10
8Individual/ml, 10
7Individual/ml, 10
6Individual/ml, 10
5Individual/ml and 10
4The D29 phage solution of individual/ml finds that the minimum bacteriophage concentration that test pen can detect is 10
6Individual/ml.
3, getting concentration respectively is 10
7Individual/ml, 10
6Individual/ml, 10
5Individual/ml, 10
4Individual/ml, 10
3Individual/ml, 10
2The shame dirt bacillus 1.0ml of individual/ml, the centrifugal supernatant that goes adds enrichment liquid (with Michaelis 7H9 nutrient culture media 7g, CaCl
22g, ampicillin 50g, amphotericin B 50g, BSA 200g, glucose 15g, oleic acid 2g and hydrogen peroxidase 0.2g are dissolved in the 1L water and prepare) 5ml, to cultivate 25 hours for 37 ℃, the centrifugal supernatant that goes adds the resuspended shame dirt of 0.9ml enrichment liquid bacillus again;
4, adding concentration in above-mentioned each sample is 10
6The bacteriophage 0.1ml of individual/ml cultivated 4 hours for 37 ℃, allowed phage-infect shame dirt bacillus to cracking;
5, above-mentioned solution is got 0.1ml detect with test pen, find when initial shame dirt bacillus number is less than 100, can't to detect bacteriophage, illustrate that this method is minimum can only detect the dirty bacillus of 100 shame.
Embodiment 4, amplification and the detection of D29 bacteriophage in tubercle bacillus
1, (sample liquid: NaOH liquid=1: 4) room temperature treatment sample to be checked is 20 minutes, the centrifugal supernatant that goes for the NaOH solution with 4%;
2, add enrichment liquid (enrichment liquid that adopts embodiment 3 to use) 5ml, to cultivate 25 hours for 37 ℃, the centrifugal supernatant that goes adds the resuspended tubercle bacillus of 0.9ml enrichment liquid again;
3, adding concentration in above-mentioned sample is 10
6The D29 phage solution 0.1ml of individual/ml, 37 ℃ of incubations 4 hours allow the phage-infect tubercle bacillus to cracking;
4, the solution after cultivating is got 0.1ml detect with test pen, if can detect bacteriophage explanation tubercle bacillus number more than 100.
Sequence table
＜110〉Inverness Medical Shanghai Co., Ltd.
＜120〉a kind of method that detects tubercle bacillus
<130>NP-09-13392
<160>4
<170>PatentIn?version?3.3
<210>1
<211>153
<212>DNA
＜213〉artificial sequence
<400>1
aagaagtgga?cccacaccct?tctggacgac?atcaccgagc?cgatcctgaa?cggcgcgaag 60
gaccagaacg?gtcgcccgct?gttcatcgag?tcgacctacg?gtgaggccgc?gagcccgttc 120
cgttcgggcc?ggatcgtcgc?ccgtccgacc?atc 153
<210>2
<211>51
<212>PRT
＜213〉artificial sequence
<400>2
KKWTHTLLDD?ITEPILNGAK?DQNGRPLFIE?STYGEAASPF?RSGRIVARPT?I 51
<210>3
<211>183
<212>DNA
＜213〉artificial sequence
<400>3
gaagcaggca?ccgctgcaca?tacgccggcc?gaactgaaga?ccatcgacct?gtccgacccg 60
tcgacctgga?ctggagccac?cggctggtcg?agcgtcggcc?acaccagccg?aggcacgctc 120
cccgagttcg?gcttcgaggg?cggcgattcc?gaggtcaagg?gctcctggca?gaagaagaag 180
ctc 183
<210>4
<211>61
<212>PRT
＜213〉artificial sequence
<400>4
EAGTAAHTPA?ELKTIDLSDP?STWTGATGWS?SVGHTSRGTL?PEFGFEGGDS?EVKGSWQKKK 60
L 61
Claims (9)
1. method that detects tubercle bacillus may further comprise the steps:
1) the tubercle bacillus incubation is cultivated with the amplification tubercle bacillus;
2) in tubercle bacillus, add bacteriophage, cracking bacterium behind the incubation;
3) with phage-resistant antibody standard gold, preparation bacteriophage colloidal gold method immune chromatograph testing pen detects bacteriophage with this test pen.
2. the method for claim 1, it is characterized in that, the antibody of colloid gold label is for discerning the antibody of tubercle bacillus bacteriophage head albumen in the test pen described in the step 3), and fixing antibody is the antibody of identification tubercle bacillus bacteriophage head albumen on the p-wire.
3. the method for claim 1, it is characterized in that, the antibody of colloid gold label is for discerning the antibody of tubercle bacillus bacteriophage tail albumen in the test pen described in the step 3), and fixing antibody is the antibody of identification tubercle bacillus bacteriophage tail albumen on the p-wire.
4. the method for claim 1, it is characterized in that, the antibody of colloid gold label is for discerning the antibody of tubercle bacillus bacteriophage head albumen in the test pen described in the step 3), and fixing antibody is the antibody of identification tubercle bacillus bacteriophage tail albumen on the p-wire.
5. the method for claim 1, it is characterized in that, the antibody of colloid gold label is for discerning the antibody of tubercle bacillus bacteriophage tail albumen in the test pen described in the step 3), and fixing antibody is the antibody of identification tubercle bacillus bacteriophage head albumen on the p-wire.
6. as each described method in the claim 2,4 and 5, it is characterized in that, the antibody specific bond tubercle bacillus bacteriophage head albumen of described identification tubercle bacillus bacteriophage head albumen, this tubercle bacillus bacteriophage head protein fragments has the amino acid sequence shown in nucleotide sequence shown in the SEQ ID NO:1 and the SEQID NO.2.
7. as each described method of claim 3-5, it is characterized in that, the antibody specific bond tubercle bacillus bacteriophage tail albumen of described identification tubercle bacillus bacteriophage tail albumen, this tubercle bacillus bacteriophage tail protein fragments has the amino acid sequence shown in nucleotide sequence shown in the SEQ ID NO:3 and the SEQ ID NO.4.
8. the method for claim 1 is characterized in that, step 1) specifically in two steps:
A) with 4% NaOH solution room temperature treatment sample to be checked 20 minutes, the centrifugal supernatant that goes;
B) add enrichment liquid, cultivated 25 hours for 37 ℃, the centrifugal supernatant that goes adds the resuspended tubercle bacillus of enrichment liquid again.
9. method as claimed in claim 8, it is characterized in that, described enrichment liquid is by Michaelis 7H9 nutrient culture media 5-7 weight portion, lime chloride 1-2 weight portion, ampicillin 25-50 weight portion, amphotericin B 25-50 weight portion, bovine serum albumin(BSA) 100-200 weight portion, glucose 10-15 weight portion, oleic acid 1-2 weight portion and hydrogen peroxidase 0.1-0.2 weight portion, all is dissolved in the pure water of 1000 weight portions being prepared from.
Priority Applications (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 200910057773 CN101995464B (en) | 2009-08-20 | 2009-08-20 | Method for detecting mycobacteria tuberculosis |
| PCT/CN2010/076121 WO2011020434A1 (en) | 2009-08-20 | 2010-08-19 | Antibody binding to the protein of the bacteriophage of mycobacterium tuberculosis, and the preparation method and use thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 200910057773 CN101995464B (en) | 2009-08-20 | 2009-08-20 | Method for detecting mycobacteria tuberculosis |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN101995464A true CN101995464A (en) | 2011-03-30 |
| CN101995464B CN101995464B (en) | 2013-08-14 |
Family
ID=43785900
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN 200910057773 Expired - Fee Related CN101995464B (en) | 2009-08-20 | 2009-08-20 | Method for detecting mycobacteria tuberculosis |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN101995464B (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104034889A (en) * | 2014-06-12 | 2014-09-10 | 江崇才 | Method for detecting mycobacterium tuberculosis |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1888076A (en) * | 2005-06-29 | 2007-01-03 | 李建林 | Fast identification method for ox tubercle bacillus and its drug sensitive test kit |
| CN101329342A (en) * | 2007-06-22 | 2008-12-24 | 王霁 | Method and reagent kit for detecting tubercle bacillus and anti-tuberculosis medicaments sensibility |
-
2009
- 2009-08-20 CN CN 200910057773 patent/CN101995464B/en not_active Expired - Fee Related
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1888076A (en) * | 2005-06-29 | 2007-01-03 | 李建林 | Fast identification method for ox tubercle bacillus and its drug sensitive test kit |
| CN101329342A (en) * | 2007-06-22 | 2008-12-24 | 王霁 | Method and reagent kit for detecting tubercle bacillus and anti-tuberculosis medicaments sensibility |
Non-Patent Citations (3)
| Title |
|---|
| TIWARI等: "Modern approaches to a rapid diagnosis of tuberculosis_ Promises and challenges ahead", 《TUBERCULOSIS》 * |
| 叶佳庆等: "噬菌体生物扩增法检测不同标本对结核病临床诊断的意义", 《实用临床医学》 * |
| 甘兵等: "噬菌体生物扩增法与涂片法在快速检测结核分支杆菌的应用比较", 《广东医学》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104034889A (en) * | 2014-06-12 | 2014-09-10 | 江崇才 | Method for detecting mycobacterium tuberculosis |
Also Published As
| Publication number | Publication date |
|---|---|
| CN101995464B (en) | 2013-08-14 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Lopez-Soriano et al. | Lateral flow immunoassay for on-site detection of Xanthomonas arboricola pv. pruni in symptomatic field samples | |
| CN102181572B (en) | Mycobacterium tuberculosis nucleic acid detection kit by utilizing RNA isothermal amplification | |
| Moreno et al. | MAb‐based competitive ELISA for the detection of antibodies against influenza D virus | |
| CN103123353A (en) | Immunodetection assay for mycobacterium tuberculosis complex | |
| WO2012163263A1 (en) | Reagents and methods for prrsv detection | |
| US10962540B2 (en) | Beta lactamase as biomarker for the specific detection of tuberculosis-complex bacteria | |
| CN106636004B (en) | TMV-CMV-PVY triple virus colloidal gold rapid detection test strip | |
| CN101363859B (en) | Test paper strip for rapidly detecting brucellosis antibody | |
| CN101709087B (en) | Puccinia triticina f.sp.tritic monoclonal antibody and application thereof | |
| RU2008130905A (en) | CHICKEN VIRUS VACCINE AND DIAGNOSTICS | |
| CN101995464B (en) | Method for detecting mycobacteria tuberculosis | |
| CN101993491B (en) | Antibody combined with tubercle bacillus bacteriophage head protein as well as preparation method and application thereof | |
| CN104185790A (en) | Devices and methods for detection of panton-valentine leukocidin (pvl) | |
| CN101993490B (en) | Antibody combined with tubercle bacillus bacteriophage tail protein as well as preparation method and application thereof | |
| CN101805723A (en) | Anti-glutamate dehydrogenase protein monoclonal antibody, preparation method thereof and use thereof | |
| CN114539393B (en) | 2019-novel coronavirus N protein single domain antibody and application thereof | |
| CN102101887B (en) | Monoclonal antibody, immune fluorescence method and kit for detecting three rusts | |
| Hannachi et al. | Enhancing the detection of Toxoplasma gondii via an anti-SAG1 scFv-alkaline phosphatase immunoconjugate | |
| Contaldo et al. | Preliminary evaluation of the use of an antiserum obtained from phytoplasma culture | |
| US8247187B2 (en) | Method for detecting pathogens | |
| CN112646007B (en) | Combined protein for detecting mycobacterium tuberculosis and detection reagent | |
| Nizkorodova et al. | REVIEW AND ANALYSIS OF ANTHRAX DIAGNOSTIC TOOLS ON THE TERRITORY OF EURASIAN ECONOMIC UNION | |
| KR20200081619A (en) | Kit for diagnosing dengue hemorrhagic fever | |
| Kulshrestha et al. | Potential uses of in vitro expressed and purified recombinant Prunus necrotic ringspot virus coat protein gene | |
| Prygiel et al. | Challenges of Diphtheria Toxin Detection |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C53 | Correction of patent of invention or patent application | ||
| CB02 | Change of applicant information |
Address after: Shanghai city 201203 libing road Zhangjiang High Tech Park of Pudong New Area No. 151 Building No. 4 floor Applicant after: ALERE (SHANGHAI) DIAGNOSTICS CO.,LTD. Address before: Shanghai city 201203 libing road Pudong New Area Zhangjiang hi tech Park No. 151 Applicant before: INVERNESS MEDICAL (SHANGHAI) Co.,Ltd. |
|
| COR | Change of bibliographic data |
Free format text: CORRECT: APPLICANT; FROM: INVERNESS MEDICAL SHANGHAI CO., LTD. TO: ALERE (SHANGHAI) DIAGNOSTICS CO., LTD. |
|
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| CP03 | Change of name, title or address | ||
| CP03 | Change of name, title or address |
Address after: 201203 1st floor, building 4, R & D building, 151 libing Road, Pudong New Area, Shanghai Patentee after: Abbott diagnostic products (Shanghai) Co.,Ltd. Address before: Shanghai city 201203 libing road Zhangjiang High Tech Park of Pudong New Area No. 151 Building No. 4 floor Patentee before: ALERE (SHANGHAI) DIAGNOSTICS Co.,Ltd. |
|
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20130814 |
|
| CF01 | Termination of patent right due to non-payment of annual fee |