CN111521820A - Application of mycobacterium tuberculosis protein in preparing product for diagnosing latent tuberculosis infected person and/or active tuberculosis - Google Patents
Application of mycobacterium tuberculosis protein in preparing product for diagnosing latent tuberculosis infected person and/or active tuberculosis Download PDFInfo
- Publication number
- CN111521820A CN111521820A CN202010371840.4A CN202010371840A CN111521820A CN 111521820 A CN111521820 A CN 111521820A CN 202010371840 A CN202010371840 A CN 202010371840A CN 111521820 A CN111521820 A CN 111521820A
- Authority
- CN
- China
- Prior art keywords
- protein
- seq
- amino acid
- ala
- val
- 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
Images
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
- G01N33/6854—Immunoglobulins
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/569—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses
- G01N33/56911—Bacteria
- G01N33/5695—Mycobacteria
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2469/00—Immunoassays for the detection of microorganisms
- G01N2469/20—Detection of antibodies in sample from host which are directed against antigens from microorganisms
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Immunology (AREA)
- Engineering & Computer Science (AREA)
- Urology & Nephrology (AREA)
- Chemical & Material Sciences (AREA)
- Biomedical Technology (AREA)
- Molecular Biology (AREA)
- Hematology (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Biotechnology (AREA)
- Pathology (AREA)
- General Physics & Mathematics (AREA)
- Food Science & Technology (AREA)
- Microbiology (AREA)
- Analytical Chemistry (AREA)
- Medicinal Chemistry (AREA)
- Cell Biology (AREA)
- Physics & Mathematics (AREA)
- Virology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Peptides Or Proteins (AREA)
Abstract
The invention provides application of a product for detecting the levels of an anti-Rv 0389 antibody, an anti-Rv 1408 antibody and an anti-Rv 2097c antibody in a sample to preparation of a product for identifying, diagnosing and/or screening active tuberculosis. The method provided by the invention can simultaneously detect the level of serum antibody response of three different mycobacterium tuberculosis proteins, so that latent tuberculosis infectors and active tuberculosis patients can be better identified and diagnosed, the sensitivity is higher, and the specificity is better.
Description
The present application is a divisional application of an invention patent application having an application date of 2016, 12 and 30 and an application number of 2016112574947.
Technical Field
The invention relates to the field of biomedicine, in particular to application of mycobacterium tuberculosis protein in preparing a product for diagnosing latent tuberculosis infectors and/or active tuberculosis.
Background
Tuberculosis caused by mycobacterium tuberculosis is still one of diseases seriously threatening human health, according to the latest report of the world health organization, 1040 ten thousands of new tuberculosis cases are found in 2015 all the world, about 180 thousands of tuberculosis cases die from tuberculosis, 91.8 thousands of new tuberculosis cases and 3.8 thousands of tuberculosis cases are found in China, and the total number of the tuberculosis cases is the fourth position in the world. The work of preventing and treating tuberculosis is still far in the world or in China, and the infection rate of tubercle bacillus is still high especially in some underdeveloped countries with poor sanitary conditions.
After a human body is infected with mycobacterium tuberculosis, about only 10% of people develop active tuberculosis, and 90% of people do not develop the active tuberculosis, but the tubercle bacillus in the human body is not completely eliminated, but is latent in the human body, namely tuberculosis latent infection, the people have no clinical symptoms, most of people do not develop the active tuberculosis in the life, and only 5% -10% of people can cause the tuberculosis when the immunity of the human body is reduced. At present, 1/3 people all over the world infect tubercle bacillus, and the number of visible latent infected people is large, so that the latent infected people need to be monitored, and when the infection state changes, the drug therapy is timely given to achieve early discovery and early treatment, so that the disease deterioration can be prevented, the treatment time and the side effect caused by the drug therapy can be reduced, and the spreading of tuberculosis can be reduced. Therefore, the detection and identification of the two states after tubercle bacillus infection are very important for the prevention and treatment of tuberculosis.
At present, the detection methods of tubercle bacillus infection comprise a plurality of methods, such as Tuberculin Skin Test (TST), gamma-interferon release test (IGRA), traditional phlegm acid-fast bacterium smear, rapid culture of mycobacteria, amplification (quantitative and qualitative) of tuberculosis specific nucleic acid fragments and the like. TST is skin allergy, has long service time, is mainly used for outpatient screening, although the test is simple and easy to implement, the same result can be produced in BCG inoculation when the tubercle bacillus infection is detected, natural infection cannot be identified, the specificity is not strong, and in addition, latent tuberculosis infection and active tuberculosis cannot be identified. The gamma interferon release assay (IGRA) is mainly used for detecting cellular immune response, although the effect of BCG vaccination can be eliminated, latent infectors and active tuberculosis patients cannot be distinguished, and treated patients and current infectors cannot be distinguished. However, the traditional sputum acid-fast bacteria smear, the rapid culture of mycobacteria and the like are mainly used for detecting active tuberculosis patients, have no significance for judging latent infection states, and in addition, bacteriological evidence may not be found for the active tuberculosis patients with atypical clinical symptoms. Therefore, the method for accurately identifying latent infection and active tuberculosis is of great value.
Based on different host protective immune responses caused by tubercle bacillus infection, the method for searching the serum marker to distinguish latent infection and active tuberculosis has good application prospect. The serological diagnosis method of antigen-antibody reaction is of great interest because of its simplicity, rapidity and easy availability of specimens, and some specific antigens of mycobacterium tuberculosis have been used for serological diagnosis of tubercle bacillus infection, such as secreted protein antigens: antigen 85 complex antigens, i.e., Ag85A, Ag85B, and Ag85C, etc.; lipoprotein antigen: 16kDa, 27kDa, 38kDa antigen, etc.; glycolipid antigens: lipoarabinomannan antigens, tuberculosyl lipoid antigens, and the like; however, in actual use, the antigens have the problems of low positive detection rate, cross immunity with other mycobacteria and the like, and although the antigens have certain values in the general survey, the monitoring of treatment effect, the judgment of prognosis and the prompt of relapse of high risk groups, the antigens cannot be used for the definite diagnosis of latent mycobacterium tuberculosis infection at present.
Therefore, it is very important to find a new method and marker capable of accurately diagnosing latent tuberculosis infectors and active tuberculosis patients.
Disclosure of Invention
The invention aims to solve the technical problems that in the prior art, products for identifying and diagnosing latent tuberculosis infectors and/or active tuberculosis have low positive detection rate and cannot identify and diagnose latent mycobacterium tuberculosis infection, and provides an application of mycobacterium tuberculosis protein in preparing products for diagnosing latent tuberculosis infectors and/or active tuberculosis.
In order to solve the technical problems, the technical scheme provided by the invention on one hand is as follows:
use of a product for detecting the level of an anti-Rv 0389 antibody, an anti-Rv 1408 antibody and/or an anti-Rv 2097c antibody in a sample in the manufacture of a product for the identification, diagnosis and/or screening of latent tuberculosis infection and/or active tuberculosis.
In the technical scheme of the invention, any product for detecting the level of the anti-Rv 0389 antibody, the anti-Rv 1408 antibody and/or the anti-Rv 2097c antibody in a sample can be used for preparing a product for identifying, diagnosing and/or screening latent tuberculosis infection and/or active tuberculosis.
Preferably, the product for detecting the level of the anti-Rv 0389 antibody, the anti-Rv 1408 antibody and/or the anti-Rv 2097c antibody in the sample comprises Rv0389 protein, Rv1408 protein and/or Rv2097c protein.
More preferably, the Rv0389 protein is a protein shown in SEQ ID No.1, or one or more amino acid residues in an amino acid sequence shown in SEQ ID No.1 are modified to have the same function as the protein shown in SEQ ID No. 1; the Rv1408 protein is shown in SEQ ID NO.2, or the protein with the same function as the protein shown in SEQ ID NO.2 after one or more amino acid residues in the amino acid sequence shown in SEQ ID NO.2 are modified; the protein Rv2097c is shown in SEQ ID NO.3, or the protein which has the same function with the protein shown in SEQ ID NO.3 after one or more amino acid residues in the amino acid sequence shown in SEQ ID NO.3 are modified.
The term "modification" as used above may be any prior art modification of proteins, including, but not limited to, substitution and/or addition and/or deletion of amino acid residues to form derivatives of the original protein.
Preferably, the detection may be any suitable detection method in the prior art, but preferably, the detection method may be an enzyme-linked immunosorbent assay, an agglutination assay, a precipitation assay, an E-rosette assay, a phagocytosis assay, an immunofluorescence assay, a colloidal gold immunochromatography assay, a spotted gold immunodiafiltration assay, and/or an electrochemiluminescence assay. More preferably, in one embodiment of the present invention, the detection method is an enzyme-linked immunosorbent assay.
Preferably, the sample may be any suitable sample obtained from the subject. Preferably, however, the sample comprises serum, plasma, saliva, urine, pleural effusion and/or cerebrospinal fluid. More preferably, in one embodiment of the present invention, the sample is serum.
Another aspect of the present invention is to provide a marker composition for identifying, diagnosing and/or screening latent tuberculosis infection and/or active tuberculosis, the marker composition consisting of an anti-Rv 0389 antibody, an anti-Rv 1408 antibody and/or an anti-Rv 2097c antibody;
wherein, the Rv0389 is a protein shown in SEQ ID NO.1, or one or more amino acid residues in the amino acid sequence shown in SEQ ID NO.1 are modified to have the same function with the protein shown in SEQ ID NO. 1; the Rv1408 is a protein shown in SEQ ID NO.2, or one or more amino acid residues in the amino acid sequence shown in SEQ ID NO.2 are modified to have the same function with the protein shown in SEQ ID NO. 2; the Rv2097c is a protein shown in SEQ ID NO.3, or one or more amino acid residues in the amino acid sequence shown in SEQ ID NO.3 are modified to have the same function with the protein shown in SEQ ID NO. 3.
In another aspect of the invention, the invention provides a use of the above marker composition in the preparation of a product for identifying, diagnosing and/or screening latent tuberculosis infection and/or active tuberculosis.
Another aspect of the present invention provides a kit for identifying, diagnosing and/or screening latent tuberculosis infection and/or active tuberculosis, the kit comprising Rv0389 protein, Rv1408 protein and/or Rv2097c protein.
Preferably, the Rv0389 protein is a protein shown in SEQ ID NO.1, or a protein which has the same function as the protein shown in SEQ ID NO.1 after one or more amino acid residues in an amino acid sequence shown in SEQ ID NO.1 are modified; the Rv1408 protein is shown in SEQ ID NO.2, or the protein with the same function as the protein shown in SEQ ID NO.2 after one or more amino acid residues in the amino acid sequence shown in SEQ ID NO.2 are modified; the protein Rv2097c is shown in SEQ ID NO.3, or the protein which has the same function with the protein shown in SEQ ID NO.3 after one or more amino acid residues in the amino acid sequence shown in SEQ ID NO.3 are modified.
The Rv0389 protein, Rv1408 protein and/or Rv2097c protein contained in the kit may be in any form, including but not limited to direct protein samples, as well as forms in which the protein is immobilized on a carrier, such as a detection chip.
The kit comprises at least kit instructions in addition to Rv0389 protein, Rv1408 protein and/or Rv2097c protein.
The specification describes criteria for determining the results: and (3) using the joint detection of the three proteins, if at least 2 antibodies in the anti-Rv 0389 antibody, the anti-Rv 1408 antibody and/or the anti-Rv 2097c antibody are positive in the detection result of the sample to be detected, determining that the sample to be detected is positive in active tuberculosis, otherwise, determining that the sample to be detected is latent tuberculosis infection.
In another aspect, the invention provides the use of the above-mentioned kit in the preparation of a product for identifying, diagnosing and/or screening latent tuberculosis infection and/or active tuberculosis.
The invention has the beneficial effects that:
the kit can simultaneously detect the level of the serum antibody response of three different mycobacterium tuberculosis proteins, thereby better identifying and diagnosing latent tuberculosis infectors and active tuberculosis patients, and having higher sensitivity and better specificity.
Drawings
FIG. 1 is a statistical chart of ELISA detection of anti-Rv 0389 antibody, anti-Rv 1408 antibody and anti-Rv 2097c antibody expression levels in sera of tuberculosis latent infected patients (n 93) and active tuberculosis patients (n 92)
FIG. 2 is a ROC graph of antibodies corresponding to the joint detection of three antigens;
DESCRIPTION OF THE SEQUENCES
SEQ ID No.1 is an amino acid sequence of the Rv0389 protein in the embodiment of the invention;
SEQ ID NO.2 is the amino acid sequence of the Rv1408 protein in the examples of the present invention;
SEQ ID NO.3 shows the amino acid sequence of the Rv2097c protein in the example of the present invention.
Detailed Description
The invention discloses application of a product for detecting the level of an anti-Rv 0389 antibody, an anti-Rv 1408 antibody and/or an anti-Rv 2097c antibody in a sample in preparing a product for identifying, diagnosing and/or screening latent tuberculosis infection and/or active tuberculosis. Those skilled in the art can modify the process parameters appropriately to achieve the desired results with reference to the disclosure herein. It is expressly intended that all such alterations and modifications which are obvious to those skilled in the art are deemed to be incorporated herein by reference, and that the techniques of the invention may be practiced and applied by those skilled in the art without departing from the spirit, scope and range of equivalents of the invention.
In the present invention, unless otherwise specified, scientific and technical terms used herein have the meanings that are commonly understood by those skilled in the art.
In order to make those skilled in the art better understand the technical solution of the present invention, the following detailed description of the present invention is provided with reference to specific embodiments.
Example 1: collection and processing of samples
The sample to be detected is serum, the collection method comprises the steps of collecting blood of a subject in an empty stomach, using a common biochemical test tube without anticoagulant, standing at room temperature, centrifuging at the normal temperature of 3000 r/min for 10 minutes after natural coagulation, collecting the serum, subpackaging in a freezing tube, and placing in a low-temperature refrigerator at the temperature of minus 80 ℃ for storage for use in experiments.
The samples used in the present invention are divided into two groups: a latent infection group and an active tuberculosis patient group; wherein, the latent infection group refers to the patients without clinical chief complaints, positive TST experiment and T-SPOT experiment, and negative chest X-ray; the active tuberculosis patient group refers to confirmed tuberculosis patients which are positive through smear or culture and do not start or just start systemic chemical drug treatment; both groups excluded diabetes, HIV infection and other autoimmune diseases.
The specimens of the latent infection group used in the invention are from epidemiological research projects of epidemiological research rooms of tuberculosis breast tumor research institute of Beijing city, the specimens of the active tuberculosis patient group are from inpatients of each disease area of tuberculosis department of Beijing thoracic hospital affiliated to the university of capital medical science, the subject is approved by the scientific research ethical committee of the Beijing thoracic hospital of capital medical university/tuberculosis breast tumor research institute of Beijing city, and all research objects agree with the conditions.
Example 2: detection of samples using ELISA assays
1. Establishing various buffers, diluents, reaction solutions and stop solutions required by an ELSIA experiment:
(1) coating buffer (pH9.6, 0.05mol/L carbonate buffer):
Na2CO31.59g，NaHCO3adding double distilled water to 1000ml for 2.93g, and adjusting the pH to 9.6;
(2) PBS solution ph 7.4:
NaCl 8.0g，KCl 0.2g，KH2PO40.2g，Na2HPO4·12H2O 2.9g，
adding double distilled water to 1000ml, and adjusting the pH value to 7.4;
(3) PBST wash ph 7.4:
adding Tween-200.5 ml into 1000ml of PBS solution, and adjusting the pH to 7.4;
(4) blocking solution (PBS solution with skim milk powder, ph 7.4):
5g of skimmed milk powder is added into 100ml of PBS solution;
(5) substrate buffer: 0.2M NaH2PO4(28.4g/L)25.7ml，
24.3ml of 0.1M citric acid (19.2g/L) and 50ml of distilled water were added.
(6) TMB (tetramethylbenzidine) use solution 0.5ml of TMB (10mg/5ml of absolute ethanol) substrate
Buffer (pH5.5)10ml 0.75% H2O232μl；
(7) Stop solution (2M H)2SO4)：21.32ml H2SO4，178.68ml H2O。
(8) Secondary antibody: horseradish peroxidase (HRP) labeled goat anti-human IgG;
(9) a 96-hole enzyme label plate; nunc Incorporated (Theromo, Gebenhagen, Denmark)
2. A detection step:
(1) coating: the Rv0389, Rv1408 and Rv2097c antigens were dissolved in coating buffer (5ug/ml) respectively; adding 100ul per well into a 96-well enzyme label plate, and standing overnight at 4 ℃;
(2) washing the plate: PBST cleaning solution 300 ul/hole, 3min, wash 3 times;
(3) and (3) sealing: blocking solution, 300 ul/hole, and incubating for 1h at room temperature;
(4) washing the plate: washing solution 300 ul/hole for 3min for 3 times;
(5) adding serum: incubating the serum to be tested at room temperature for 1h at 100 ul/hole after dilution at a ratio of 1: 400;
(6) washing the plate: washing solution 300 ul/hole for 3min for 5 times;
(7) adding an enzyme-labeled secondary antibody: 100ul of freshly diluted enzyme-labeled antibody (1:30000) per well, and incubating for 1h at room temperature;
(8) washing the plate: washing solution 300 ul/hole for 3min for 5 times;
(9) color development: incubation is carried out for 5-10min at room temperature in a dark place with 100 ul/hole of TMB developing solution prepared freshly;
(10) and (4) terminating: stop solution 2M H2SO450 ul/well;
(11) and (3) determination: the microplate reader was adjusted to 450nm and the OD of each well was measured after zeroing with the first blank PBS control well.
3. Results analysis and criteria for determination:
using SPSS22.0 software to draw an ROC curve, obtaining specificity and sensitivity corresponding to different critical points according to coordinate values, then calculating a Yoden index (the Yoden index is specificity + sensitivity-1), and obtaining the OD values of the critical points of the three antigens respectively, wherein the OD values of the critical points are that Rv0389 is 0.291, that of Rv1408 is 0.283, and that of Rv2097c is 0.308;
the results are shown in figure 1, and show that the three proteins have the function of distinguishing latent tuberculosis infection and active tuberculosis.
And (3) judging standard:
and (3) using the joint detection of the three proteins, if at least 2 antibodies in the anti-Rv 0389 antibody, the anti-Rv 1408 antibody and/or the anti-Rv 2097c antibody are positive in the detection result of the sample to be detected, determining that the sample to be detected is positive in active tuberculosis, otherwise, determining that the sample to be detected is latent tuberculosis infection.
Example 3: specificity and sensitivity of method for differential diagnosis of active tuberculosis patients and latent infected patients by applying ELISA
Adopting 92 parts of serum of a patient clinically diagnosed with active tuberculosis and 93 parts of serum of a latent infected patient; the ELISA method of examples 1 and 2 was used to determine whether the test subject was positive for active tuberculosis by the criteria of example 2, and the results are shown in FIG. 2, with assay specificity of 86.0% and sensitivity of 71.7%. In FIG. 2, the abscissa 1-specificity represents the false positive rate, and the ordinate sensitivity represents the true positive rate.
The results prove that the kit can detect the level of the serum antibody response of three different mycobacterium tuberculosis proteins, can better identify and diagnose latent tuberculosis infectors and active tuberculosis patients, and has higher sensitivity and better specificity.
The foregoing is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, various modifications and decorations can be made without departing from the principle of the present invention, and these modifications and decorations should also be regarded as the protection scope of the present invention.
Sequence listing
<110> Beijing thoracic Hospital affiliated to capital medical university
<120> use of protein of mycobacterium tuberculosis in preparing products for diagnosing latent tuberculosis infection and/or active tuberculosis
<130>None
<160>3
<170>SIPOSequenceListing 1.0
<210>1
<211>419
<212>PRT
<213>Bacteria
<400>1
Val Ile Asp Gly Trp Thr Glu Glu Gln His Glu Pro Thr Val Arg His
1 5 10 15
Glu Arg Pro Ala Ala Pro Gln Asp Val Arg Arg Val Met Leu Leu Gly
20 25 30
Ser Ala Glu Pro Ser Arg Glu Leu Ala Ile Ala Leu Gln Gly Leu Gly
35 40 45
Ala Glu Val Ile Ala Val Asp Gly Tyr Val Gly Ala Pro Ala His Arg
50 55 60
Ile Ala Asp Gln Ser Val Val Val Thr Met Thr Asp Ala Glu Glu Leu
65 70 75 80
Thr Ala Val Ile Arg Arg Leu Gln Pro Asp Phe Leu Val Thr Val Thr
85 90 95
Ala Ala Val Ser Val Asp Ala Leu Asp Ala Val Glu Gln Ala Asp Gly
100 105 110
Glu Cys Thr Glu Leu Val Pro Asn Ala Arg Ala Val Arg Cys Thr Ala
115 120 125
Asp Arg Glu Gly Leu Arg Arg Leu Ala Ala Asp Gln Leu Gly Leu Pro
130 135 140
Thr Ala Pro Phe Trp Phe Val Gly Ser Leu Gly Glu Leu Gln Ala Val
145 150 155 160
Ala Val His Ala Gly Phe Pro Leu Leu Val Ser Pro Val Ala Gly Val
165 170 175
Ala Gly Gln Gly Ser Ser Val Val Ala Gly Pro Asn Glu Val Glu Pro
180 185 190
Ala Trp Gln Arg Ala Ala Gly His Gln Val Gln Pro Gln Thr Gly Gly
195 200 205
Val Ser Pro Arg Val Cys Ala Glu Ser Val Val Glu Ile Glu Phe Leu
210 215 220
Val Thr Met Ile Val Val Cys Ser Gln Gly Pro Asn Gly Pro Leu Ile
225 230 235 240
Glu Phe Cys Ala Pro Ile Gly His Arg Asp Ala Asp Ala Gly Glu Leu
245 250 255
Glu Ser Trp Gln Pro Gln Lys Leu Ser Thr Ala Ala Leu Asp Ala Ala
260 265 270
Lys Ser Ile Ala Ala Arg Ile Val Lys Ala Leu Gly Gly Arg Gly Val
275 280 285
Phe Gly Val Glu Leu Met Ile Asn Gly Asp Glu Val Tyr Phe Ala Asp
290 295 300
Val Thr Val Cys Pro Ala Gly Ser Ala Trp Val Thr Val Arg Ser Gln
305 310 315 320
Arg Leu Ser Val Phe Glu Leu Gln Ala Arg Ala Ile Leu Gly Leu Ala
325 330 335
Val Asp Thr Leu Met Ile Ser Pro Gly Ala Ala Arg Val Ile Asn Pro
340 345 350
Asp His Thr Ala Gly Arg Ala Ala Val Gly Ala Ala Pro Pro Ala Asp
355 360 365
Ala Leu Thr Gly Ala Leu Gly Val Pro Glu Ser Asp Val Val Ile Phe
370 375 380
Gly Arg Gly Leu Gly Val Ala Leu Ala Thr Ala Pro Glu Val Ala Ile
385 390 395 400
Ala Arg Glu Arg Ala Arg Glu Val Ala Ser Arg Leu Asn Val Pro Asp
405 410 415
Ser Arg Glu
<210>2
<211>232
<212>PRT
<213>Bacteria
<400>2
Val Ser Leu Met Ala Gly Ser Thr Gly Gly Pro Leu Ile Ala Pro Ser
1 5 10 15
Ile Leu Ala Ala Asp Phe Ala Arg Leu Ala Asp Glu Ala Ala Ala Val
20 25 30
Asn Gly Ala Asp Trp Leu His Val Asp Val Met Asp Gly His Phe Val
35 40 45
Pro Asn Leu Thr Ile Gly Leu Pro Val Val Glu Ser Leu Leu Ala Val
50 55 60
Thr Asp Ile Pro Met Asp Cys His Leu Met Ile Asp Asn Pro Asp Arg
65 7075 80
Trp Ala Pro Pro Tyr Ala Glu Ala Gly Ala Tyr Asn Val Thr Phe His
85 90 95
Ala Glu Ala Thr Asp Asn Pro Val Gly Val Ala Arg Asp Ile Arg Ala
100 105 110
Ala Gly Ala Lys Ala Gly Ile Ser Val Lys Pro Gly Thr Pro Leu Glu
115 120 125
Pro Tyr Leu Asp Ile Leu Pro His Phe Asp Thr Leu Leu Val Met Ser
130 135 140
Val Glu Pro Gly Phe Gly Gly Gln Arg Phe Ile Pro Glu Val Leu Ser
145 150 155 160
Lys Val Arg Ala Val Arg Lys Met Val Asp Ala Gly Glu Leu Thr Ile
165 170 175
Leu Val Glu Ile Asp Gly Gly Ile Asn Asp Asp Thr Ile Glu Gln Ala
180 185 190
Ala Glu Ala Gly Val Asp Cys Phe Val Ala Gly Ser Ala Val Tyr Gly
195 200 205
Ala Asp Asp Pro Ala Ala Ala Val Ala Ala Leu Arg Arg Gln Ala Gly
210 215 220
Ala Ala Ser Leu His Leu Ser Leu
225 230
<210>3
<211>452
<212>PRT
<213>Bacteria
<400>3
Val Gln Arg Arg Ile Met Gly Ile Glu Thr Glu Phe Gly Val Thr Cys
1 5 10 15
Thr Phe His Gly His Arg Arg Leu Ser Pro Asp Glu Val Ala Arg Tyr
20 25 30
Leu Phe Arg Arg Val Val Ser Trp Gly Arg Ser Ser Asn Val Phe Leu
35 40 45
Arg Asn Gly Ala Arg Leu Tyr Leu Asp Val Gly Ser His Pro Glu Tyr
50 55 60
Ala Thr Ala Glu Cys Asp Ser Leu Val Gln Leu Val Thr His Asp Arg
65 70 75 80
Ala Gly Glu Trp Val Leu Glu Asp Leu Leu Val Asp Ala Glu Gln Arg
85 90 95
Leu Ala Asp Glu Gly Ile Gly Gly Asp Ile Tyr Leu Phe Lys Asn Asn
100 105 110
Thr Asp Ser Ala Gly Asn Ser Tyr Gly Cys His Glu Asn Tyr Leu Ile
115 120 125
Val Arg Ala Gly Glu Phe Ser Arg Ile Ser Asp Val Leu Leu Pro Phe
130 135 140
Leu Val Thr Arg Gln Leu Ile Cys Gly Ala Gly Lys Val Leu Gln Thr
145 150 155 160
Pro Lys Ala Ala Thr Tyr Cys Leu Ser Gln Arg Ala Glu His Ile Trp
165 170 175
Glu Gly Val Ser Ser Ala Thr Thr Arg Ser Arg Pro Ile Ile Asn Thr
180 185 190
Arg Asp Glu Pro His Ala Asp Ala Glu Lys Tyr Arg Arg Leu His Val
195 200 205
Ile Val Gly Asp Ser Asn Met Ser Glu Thr Thr Thr Met Leu Lys Val
210 215 220
Gly Thr Ala Ala Leu Val Leu Glu Met Ile Glu Ser Gly Val Ala Phe
225 230 235 240
Arg Asp Phe Ser Leu Asp Asn Pro Ile Arg Ala Ile Arg Glu Val Ser
245 250 255
His Asp Val Thr Gly Arg Arg Pro Val Arg Leu Ala Gly Gly Arg Gln
260 265 270
Ala Ser Ala Leu Asp Ile Gln Arg Glu Tyr Tyr Thr Arg Ala Val Glu
275 280 285
His Leu Gln Thr Arg Glu Pro Asn Ala Gln Ile Glu Gln Val Val Asp
290 295300
Leu Trp Gly Arg Gln Leu Asp Ala Val Glu Ser Gln Asp Phe Ala Lys
305 310 315 320
Val Asp Thr Glu Ile Asp Trp Val Ile Lys Arg Lys Leu Phe Gln Arg
325 330 335
Tyr Gln Asp Arg Tyr Asp Met Glu Leu Ser His Pro Lys Ile Ala Gln
340 345 350
Leu Asp Leu Ala Tyr His Asp Ile Lys Arg Gly Arg Gly Ile Phe Asp
355 360 365
Leu Leu Gln Arg Lys Gly Leu Ala Ala Arg Val Thr Thr Asp Glu Glu
370 375 380
Ile Ala Glu Ala Val Asp Gln Pro Pro Gln Thr Thr Arg Ala Arg Leu
385 390 395 400
Arg Gly Glu Phe Ile Ser Ala Ala Gln Glu Ala Gly Arg Asp Phe Thr
405 410 415
Val Asp Trp Val His Leu Lys Leu Asn Asp Gln Ala Gln Arg Thr Val
420 425 430
Leu Cys Lys Asp Pro Phe Arg Ala Val Asp Glu Arg Val Lys Arg Leu
435 440 445
Ile Ala Ser Met
450
Claims (10)
1. Use of a product for detecting the levels of an anti-Rv 0389 antibody, an anti-Rv 1408 antibody and an anti-Rv 2097c antibody in a sample for the manufacture of a product for the identification, diagnosis and/or screening of active tuberculosis.
2. The use of claim 1, wherein the product for detecting the levels of anti-Rv 0389, anti-Rv 1408 and anti-Rv 2097c antibodies in a sample comprises Rv0389, Rv1408 and Rv2097c protein.
3. The use according to claim 2, wherein the Rv0389 protein is a protein shown in SEQ ID No.1, or a protein with one or more amino acid residues in the amino acid sequence shown in SEQ ID No.1 modified to have the same function as the protein shown in SEQ ID No. 1; the Rv1408 protein is shown in SEQ ID NO.2, or the protein with the same function as the protein shown in SEQ ID NO.2 after one or more amino acid residues in the amino acid sequence shown in SEQ ID NO.2 are modified; the protein Rv2097c is shown in SEQ ID NO.3, or the protein which has the same function with the protein shown in SEQ ID NO.3 after one or more amino acid residues in the amino acid sequence shown in SEQ ID NO.3 are modified.
4. Use according to any one of claims 1 to 3, wherein the detection method comprises an enzyme-linked immunosorbent assay, an agglutination assay, a precipitation assay, an E rosette assay, a phagocytosis assay, an immunofluorescence assay, a colloidal gold immunochromatography assay, a dot immunogold diafiltration assay and/or an electrochemiluminescence assay.
5. The use according to any one of claims 1 to 3, wherein the sample comprises serum, plasma, saliva, urine, pleural effusion and/or cerebrospinal fluid.
6. A marker composition for use in the identification, diagnosis and/or screening of active tuberculosis, wherein said marker composition consists of an anti-Rv 0389 antibody, an anti-Rv 1408 antibody and an anti-Rv 2097c antibody;
wherein, the Rv0389 is a protein shown in SEQ ID NO.1, or one or more amino acid residues in the amino acid sequence shown in SEQ ID NO.1 are modified to have the same function with the protein shown in SEQ ID NO. 1; the Rv1408 is a protein shown in SEQ ID NO.2, or one or more amino acid residues in the amino acid sequence shown in SEQ ID NO.2 are modified to have the same function with the protein shown in SEQ ID NO. 2; the Rv2097c is a protein shown in SEQ ID NO.3, or one or more amino acid residues in the amino acid sequence shown in SEQ ID NO.3 are modified to have the same function with the protein shown in SEQ ID NO. 3.
7. Use of a marker composition according to claim 6 in the manufacture of a product for the identification, diagnosis and/or screening of active tuberculosis.
8. A kit for identifying, diagnosing and/or screening active tuberculosis, characterized in that it comprises Rv0389 protein, Rv1408 protein and Rv2097c protein.
9. The kit according to claim 8, wherein the Rv0389 protein is a protein shown in SEQ ID No.1, or a protein with the same function as the protein shown in SEQ ID No.1 after one or more amino acid residues in the amino acid sequence shown in SEQ ID No.1 are modified; the Rv1408 protein is shown in SEQ ID NO.2, or the protein with the same function as the protein shown in SEQ ID NO.2 after one or more amino acid residues in the amino acid sequence shown in SEQ ID NO.2 are modified; the protein Rv2097c is shown in SEQ ID NO.3, or the protein which has the same function with the protein shown in SEQ ID NO.3 after one or more amino acid residues in the amino acid sequence shown in SEQ ID NO.3 are modified.
10. Use of a kit according to claim 8 or 9 for the manufacture of a product for the identification, diagnosis and/or screening of active tuberculosis.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010371840.4A CN111521820B (en) | 2016-12-30 | 2016-12-30 | Use of mycobacterium tuberculosis proteins in the preparation of a product for diagnosing tuberculosis latency infected persons and/or active tuberculosis |
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010371840.4A CN111521820B (en) | 2016-12-30 | 2016-12-30 | Use of mycobacterium tuberculosis proteins in the preparation of a product for diagnosing tuberculosis latency infected persons and/or active tuberculosis |
| CN201611257494.7A CN108267588B (en) | 2016-12-30 | 2016-12-30 | Application of mycobacterium tuberculosis protein in preparing product for diagnosing latent tuberculosis infected person and/or active tuberculosis |
Related Parent Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201611257494.7A Division CN108267588B (en) | 2016-12-30 | 2016-12-30 | Application of mycobacterium tuberculosis protein in preparing product for diagnosing latent tuberculosis infected person and/or active tuberculosis |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN111521820A true CN111521820A (en) | 2020-08-11 |
| CN111521820B CN111521820B (en) | 2023-06-16 |
Family
ID=62754583
Family Applications (3)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202010371676.7A Active CN111551742B (en) | 2016-12-30 | 2016-12-30 | Use of mycobacterium tuberculosis proteins in the preparation of a product for diagnosing tuberculosis latency infected persons and/or active tuberculosis |
| CN202010371840.4A Active CN111521820B (en) | 2016-12-30 | 2016-12-30 | Use of mycobacterium tuberculosis proteins in the preparation of a product for diagnosing tuberculosis latency infected persons and/or active tuberculosis |
| CN201611257494.7A Active CN108267588B (en) | 2016-12-30 | 2016-12-30 | Application of mycobacterium tuberculosis protein in preparing product for diagnosing latent tuberculosis infected person and/or active tuberculosis |
Family Applications Before (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202010371676.7A Active CN111551742B (en) | 2016-12-30 | 2016-12-30 | Use of mycobacterium tuberculosis proteins in the preparation of a product for diagnosing tuberculosis latency infected persons and/or active tuberculosis |
Family Applications After (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201611257494.7A Active CN108267588B (en) | 2016-12-30 | 2016-12-30 | Application of mycobacterium tuberculosis protein in preparing product for diagnosing latent tuberculosis infected person and/or active tuberculosis |
Country Status (1)
| Country | Link |
|---|---|
| CN (3) | CN111551742B (en) |
Citations (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2003004520A2 (en) * | 2001-07-04 | 2003-01-16 | Health Protection Agency | Mycobacterial antigens expressed during latency |
| US20040213776A1 (en) * | 2002-12-06 | 2004-10-28 | Darwin Katerina Heran | Mycobacterium tuberculosis (MTB) targets for antibiotic therapy |
| US20060182685A1 (en) * | 2004-09-04 | 2006-08-17 | Bishai William R | Hollow fiber technique for in vivo study of cell populations |
| US20060228712A1 (en) * | 1999-12-16 | 2006-10-12 | Kyowa Hakko Kogyo Co., Ltd. | Novel polynucleotides |
| CN101365714A (en) * | 2005-07-01 | 2009-02-11 | 福赛特儿童口腔医院 | Tuberculosis antigen detection assays and vaccines |
| CN102590502A (en) * | 2012-01-11 | 2012-07-18 | 北京市结核病胸部肿瘤研究所 | Kit for auxiliary diagnosis of tuberculosis patients |
| US20120283118A1 (en) * | 2011-05-06 | 2012-11-08 | Alito Alicia Elsa | Methods for detecting mycobacterium tuberculosis antigens |
| CN103698531A (en) * | 2013-11-25 | 2014-04-02 | 广东体必康生物科技有限公司 | Application of Rv1860, Rv0173 and/or Rv1812c protein in preparation of products used for diagnosis of active tuberculosis |
| CN103698511A (en) * | 2013-11-25 | 2014-04-02 | 广东体必康生物科技有限公司 | Application of mycobacterium tuberculosis protein in preparation of products used for diagnosis of latent tuberculosis infection |
| CN104805063A (en) * | 2014-01-24 | 2015-07-29 | 中国人民解放军第三〇九医院 | Mycobacterium tuberculosis latent infection related proteins, preparation and applications thereof |
| CN105588944A (en) * | 2016-02-25 | 2016-05-18 | 首都医科大学附属北京胸科医院 | Application of AGP1, SERPINA3 and CDH1 content detection system in screening active tuberculosis patients |
Family Cites Families (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| AU2012202486B2 (en) * | 1999-07-13 | 2014-08-14 | Statens Serum Institut | Tuberculosis vaccine and diagnostics based on the Mycobacterium tuberculosis esat-6 gene family |
| NZ562142A (en) * | 2005-03-31 | 2009-12-24 | Univ Leiden Medical Ct | Methods and means for diagnostics, prevention and treatment of Mycobacterium infections and tuberculosis disease |
| WO2007131291A1 (en) * | 2006-05-16 | 2007-11-22 | Proteome Systems Limited | Methods of diagnosis and treatment of m. tuberculosis infection and reagents therefore |
| CN101294964B (en) * | 2007-11-27 | 2012-06-27 | 复旦大学附属华山医院 | Reagent and method for detecting active tuberculosis and tuberculosis dormant infection |
| GB0906215D0 (en) * | 2009-04-09 | 2009-05-20 | Lalvani Ajit | Diagnostic test |
| CN103698530B (en) * | 2013-11-25 | 2015-08-19 | 广东体必康生物科技有限公司 | The purposes of Mycobacterium tuberculosis albumen in the phthisical product of preparation diagnostic activities |
-
2016
- 2016-12-30 CN CN202010371676.7A patent/CN111551742B/en active Active
- 2016-12-30 CN CN202010371840.4A patent/CN111521820B/en active Active
- 2016-12-30 CN CN201611257494.7A patent/CN108267588B/en active Active
Patent Citations (12)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20060228712A1 (en) * | 1999-12-16 | 2006-10-12 | Kyowa Hakko Kogyo Co., Ltd. | Novel polynucleotides |
| WO2003004520A2 (en) * | 2001-07-04 | 2003-01-16 | Health Protection Agency | Mycobacterial antigens expressed during latency |
| US20040241826A1 (en) * | 2001-07-04 | 2004-12-02 | James Brian William | Mycobacterial antigens expressed during latency |
| US20040213776A1 (en) * | 2002-12-06 | 2004-10-28 | Darwin Katerina Heran | Mycobacterium tuberculosis (MTB) targets for antibiotic therapy |
| US20060182685A1 (en) * | 2004-09-04 | 2006-08-17 | Bishai William R | Hollow fiber technique for in vivo study of cell populations |
| CN101365714A (en) * | 2005-07-01 | 2009-02-11 | 福赛特儿童口腔医院 | Tuberculosis antigen detection assays and vaccines |
| US20120283118A1 (en) * | 2011-05-06 | 2012-11-08 | Alito Alicia Elsa | Methods for detecting mycobacterium tuberculosis antigens |
| CN102590502A (en) * | 2012-01-11 | 2012-07-18 | 北京市结核病胸部肿瘤研究所 | Kit for auxiliary diagnosis of tuberculosis patients |
| CN103698531A (en) * | 2013-11-25 | 2014-04-02 | 广东体必康生物科技有限公司 | Application of Rv1860, Rv0173 and/or Rv1812c protein in preparation of products used for diagnosis of active tuberculosis |
| CN103698511A (en) * | 2013-11-25 | 2014-04-02 | 广东体必康生物科技有限公司 | Application of mycobacterium tuberculosis protein in preparation of products used for diagnosis of latent tuberculosis infection |
| CN104805063A (en) * | 2014-01-24 | 2015-07-29 | 中国人民解放军第三〇九医院 | Mycobacterium tuberculosis latent infection related proteins, preparation and applications thereof |
| CN105588944A (en) * | 2016-02-25 | 2016-05-18 | 首都医科大学附属北京胸科医院 | Application of AGP1, SERPINA3 and CDH1 content detection system in screening active tuberculosis patients |
Non-Patent Citations (2)
| Title |
|---|
| 罗开军等: "结核分枝杆菌休眠期和活跃期差异表达基因分析", 《中国病原生物学杂志》 * |
| 罗开军等: "结核分枝杆菌休眠期和活跃期差异表达基因分析", 《中国病原生物学杂志》, no. 12, 30 December 2010 (2010-12-30), pages 889 - 894 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN111551742B (en) | 2023-06-16 |
| CN108267588B (en) | 2020-10-23 |
| CN108267588A (en) | 2018-07-10 |
| CN111551742A (en) | 2020-08-18 |
| CN111521820B (en) | 2023-06-16 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| JP2011095275A (en) | Detection of tuberculosis and infection by mycobacterium tuberculosis using hbha | |
| MXPA01004469A (en) | Tuberculosis diagnostic test. | |
| JP2019015737A (en) | Immunological detection method and kit for Mycoplasma pneumoniae | |
| CN106939035B (en) | Mycobacterium tuberculosis T cell epitope polypeptide and application thereof | |
| CN108267589B (en) | Application of mycobacterium tuberculosis protein in preparing product for diagnosing latent tuberculosis infected person and/or active tuberculosis | |
| KR101775851B1 (en) | Method and kit for diagnosis of tuberculosis based on Mycobacterium tuberculosis antigen-specific antibody reaction | |
| McConkey S et al. | Evaluation of a rapid-format antibody test and the tuberculin skin test for diagnosis of tuberculosis in two contrasting endemic settings | |
| CN103063837B (en) | Reagent, method and kit for detecting mycobacterial infection | |
| JP2019508684A (en) | Methods and kits for the diagnosis of active tuberculosis | |
| CN108267588B (en) | Application of mycobacterium tuberculosis protein in preparing product for diagnosing latent tuberculosis infected person and/or active tuberculosis | |
| Abeijon et al. | Novel antigen detection assay to monitor therapeutic efficacy of visceral leishmaniasis | |
| Xifeng et al. | Development and evaluation of a colloidal gold immunochromatographic assay based on recombinant protein CatL1D for serodiagnosis of sheep fasciolosis | |
| Harinath et al. | A cocktail of affinity-purified antibodies reactive with diagnostically useful mycobacterial antigens ES-31, ES-43, and EST-6 for detecting the presence of Mycobacterium tuberculosis | |
| KR102132964B1 (en) | Method for Diagnosing Scrub Typhus Using Exosome derived from Orientia tsutsugamushi | |
| Sarkari et al. | A comparative study of antigen and antibody detection in visceral leishmaniasis using serum and urine-based ELISA | |
| CN109061166B (en) | ELISpot detection kit for detecting bovine tuberculosis | |
| Tran et al. | Clinical Utility of Combined Whole-cell Antigen and Recombinant Hemolysis Co-regulated Protein 1-Enzyme-linked Immunosorbent Assays Reveals Underdiagnosed Cases of Melioidosis in Vietnam | |
| Nazri et al. | Detection of Plasmodium knowlesi in whole blood samples with sandwich enzyme-linked immunosorbent assay (ELISA) using rhoptry-associated protein 1 specific polyclonal antibodies | |
| CN106814194B (en) | A kind of kit for distinguishing activity and latent tuberculosis infects | |
| Shende et al. | A low molecular weight ES-20 protein released in vivo and in vitro with diagnostic potential in lymph node tuberculosis | |
| CN116400077A (en) | Antigen composition for distinguishing active tuberculosis and latent tuberculosis and application thereof | |
| KR20160139888A (en) | Method and kit for diagnosis of tuberculosis based on Mycobacterium tuberculosis antigen-specific antibody reaction | |
| KR101135668B1 (en) | Composition for diagnosis of M. Tuberculosis and Method for diagnosis using the same | |
| Meshram et al. | Sandwich elisa for tubercular antigen detection in AFB sputum positive tubercular patients in rural based Tertiary Care Hospital | |
| Majumdar et al. | A prospective study of inhouse developed SEVA TB ELISA using cocktail of antigens and their immunoglobulins in the diagnosis of tuberculosis suspected patients in a tertiary care hospital located in rural area |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |