CN111551742B - Use of mycobacterium tuberculosis protein in preparing products for diagnosing tuberculosis latent infection and/or active pulmonary tuberculosis - Google Patents

Abstract

The invention provides the use of the product for detecting the levels of anti-Rv0389 antibody, anti-Rv1408 antibody and anti-Rv2097c antibody in samples in the preparation of products for identification, diagnosis and/or screening of tuberculosis latent infection. The method provided by the invention can detect the serum antibody response levels of three different mycobacterium tuberculosis proteins at the same time, so as to better differentiate and diagnose latent tuberculosis infection patients and active tuberculosis patients, with higher sensitivity and better specificity.

Description

Mycobacterium tuberculosis proteins in the preparation of diagnostic latent tuberculosis infection and/or active Uses in tuberculosis products

This application is a divisional application of an invention patent application with an application date of December 30, 2016 and an application number of 2016112574947.

technical field

The invention relates to the field of biomedicine, in particular to the use of mycobacterium tuberculosis protein in preparing products for diagnosing tuberculosis latent infection and/or active pulmonary tuberculosis.

Background technique

After the human body is infected with Mycobacterium tuberculosis, only about 10% of people develop active tuberculosis, while 90% of people do not get sick, but the tuberculosis bacteria in the body are not completely eliminated, but dormant in the human body, called tuberculosis Latent infection, these people do not have any clinical symptoms, and most people do not get sick in their lifetime, and only 5%--10% people can cause the generation of tuberculosis when body immunity declines. At present, about 1/3 of the people in the world are infected with tuberculosis. It can be seen that there are a large number of latent infections. Therefore, it is necessary to monitor the latent infections and give timely drug treatment when the infection status changes, so as to achieve early detection and early detection. Treatment, which can not only prevent the deterioration of the disease, reduce the treatment time and side effects caused by drug treatment, but also reduce the spread of tuberculosis. Therefore, the detection and identification of the two states after tuberculosis infection has a very important effect on the prevention and treatment of tuberculosis.

At present, there are many detection methods for tuberculosis infection, such as tuberculin skin test (TST), interferon-gamma release assay (IGRA), traditional sputum smear of acid-fast bacteria, rapid culture of mycobacteria, and tuberculosis-specific nucleic acid fragments. Amplification (quantitative and qualitative), etc. TST is a skin allergy and has been used for a long time. It is mainly used for outpatient screening. Although the test is simple and easy, when detecting tuberculosis infection, BCG vaccination will produce the same results, and natural infection cannot be identified, and the specificity is not strong. In addition, latent tuberculosis infection cannot be distinguished from active tuberculosis. The gamma-interferon release assay (IGRA) is mainly used to detect cellular immune responses. Although it can rule out the impact of BCG vaccination, it cannot distinguish latent infection from active tuberculosis patients, let alone distinguish treated patients from current infection. However, traditional sputum smears of acid-fast bacteria and rapid culture of mycobacteria are mainly used to detect active tuberculosis patients, which are meaningless for the judgment of latent infection status. In addition, for active tuberculosis patients with atypical clinical symptoms, bacteria may not be found scientific evidence. Therefore, it is of great value to find a method that can accurately identify latent infection and active tuberculosis.

Based on the differences in host protective immune responses after tuberculosis infection, finding serum markers to distinguish latent infection from active tuberculosis has a good application prospect. The serological diagnosis method of antigen-antibody reaction has attracted much attention because of its simplicity, rapidity, and easy access to samples. Now some specific antigens of Mycobacterium tuberculosis have been used for serological diagnosis of tuberculosis infection, such as Secreted protein antigens: antigen 85 complex antigens, namely Ag85A, Ag85B and Ag85C, etc.; lipoprotein antigens: 16kDa, 27kDa, 38kDa antigens, etc.; glycolipid antigens: lipoarabinomannan antigens, tuberculosis carbohydrate lipids Antigens, etc.; however, in actual use, these antigens have low positive detection rates and cross-immunity with other mycobacteria. However, it still cannot be used for the diagnosis of latent Mycobacterium tuberculosis infection.

Therefore, it is extremely important to find a new method and marker that can accurately diagnose latent tuberculosis infection and active tuberculosis patients.

Contents of the invention

The technical problem to be solved by the present invention is aimed at the low positive detection rate of products for differential diagnosis of latent tuberculosis infection and/or active pulmonary tuberculosis in the prior art, and the problem that latent Mycobacterium tuberculosis infection cannot be differentially diagnosed. Use of a Mycobacterium tuberculosis protein in preparing products for diagnosing tuberculosis latent infection and/or active pulmonary tuberculosis.

In order to solve the above technical problems, the technical solution provided by the present invention on the one hand is:

Use of the product for detecting the level of anti-Rv0389 antibody, anti-Rv1408 antibody and/or anti-Rv2097c antibody in a sample in the preparation of products for identifying, diagnosing and/or screening tuberculosis latent infection and/or active pulmonary tuberculosis.

In the technical scheme of the present invention, any product that detects the level of anti-Rv0389 antibody, anti-Rv1408 antibody and/or anti-Rv2097c antibody in the sample can be used for the preparation of identification, diagnosis and/or screening for tuberculosis latent infection and/or active pulmonary tuberculosis The product.

Preferably, the above-mentioned product for detecting the level of anti-Rv0389 antibody, anti-Rv1408 antibody and/or anti-Rv2097c antibody in the sample contains Rv0389 protein, Rv1408 protein and/or Rv2097c protein.

More preferably, the Rv0389 protein is shown in SEQ ID NO.1, or one or several amino acid residues in the amino acid sequence shown in SEQ ID NO.1 are engineered to have the same A protein with the same function; the Rv1408 protein is shown in SEQ ID NO.2, or one or several amino acid residues in the amino acid sequence shown in SEQ ID NO.2 are modified and the protein shown in SEQ ID NO.2 A protein with the same function; the Rv2097c protein is shown in SEQ ID NO.3, or one or several amino acid residues in the amino acid sequence shown in SEQ ID NO.3 are modified and shown in SEQ ID NO.3 protein A protein with the same function.

The above-mentioned term "modification" can use any protein modification method in the prior art, including but not limited to, substitution and/or addition and/or deletion of amino acid residues, so as to form derivatives of the original protein.

Preferably, the detection can be any suitable detection method in the prior art, but as a preference, the detection method can be enzyme-linked immunosorbent assay, agglutination test, precipitation test, E rosette test, small phagocytosis test, immune Fluorescence experiments, colloidal gold immunochromatography experiments, spot gold immunofiltration experiments and/or electrochemiluminescence experiments. More preferably, in one embodiment of the present invention, the detection method is enzyme-linked immunosorbent assay.

Preferably, the sample can be any suitable sample obtained from the object to be detected. But preferably, the sample includes serum, plasma, saliva, urine, ascites and/or cerebrospinal fluid. More preferably, in one embodiment of the present invention, the sample is serum.

Another aspect of the present invention provides a marker composition for identifying, diagnosing and/or screening tuberculosis latent infection and/or active pulmonary tuberculosis, said marker composition consisting of anti-Rv0389 antibody, anti-Rv1408 antibody and/or anti-Rv2097c antibody composition;

Wherein, the Rv0389 is shown in SEQ ID NO.1, or one or several amino acid residues in the 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 Protein; the Rv1408 is shown in SEQ ID NO.2, or one or several amino acid residues in the amino acid sequence shown in SEQ ID NO.2 are modified to have the same function as the protein shown in SEQ ID NO.2 Protein; the Rv2097c is shown in SEQ ID NO.3, or one or several amino acid residues in the amino acid sequence shown in SEQ ID NO.3 are modified to have the same function as the protein shown in SEQ ID NO.3 protein.

Another aspect of the present invention is to provide a use of the above marker composition in the preparation of products for identifying, diagnosing and/or screening tuberculosis latent infection and/or active tuberculosis.

Another aspect of the present invention is to provide a kit for identifying, diagnosing and/or screening tuberculosis latent infection and/or active pulmonary tuberculosis, said kit comprising Rv0389 protein, Rv1408 protein and/or Rv2097c protein.

Preferably, the Rv0389 protein is shown in SEQ ID NO.1, or one or several amino acid residues in the amino acid sequence shown in SEQ ID NO.1 are modified to have the same protein as the protein shown in SEQ ID NO.1 Functional protein; the Rv1408 protein is shown in SEQ ID NO.2, or one or several amino acid residues in the amino acid sequence shown in SEQ ID NO.2 are modified to have the protein shown in SEQ ID NO.2 A protein with the same function; the Rv2097c protein is shown in SEQ ID NO.3, or one or several amino acid residues in the amino acid sequence shown in SEQ ID NO.3 are modified and the protein shown in SEQ ID NO.3 proteins with the same function.

The Rv0389 protein, Rv1408 protein and/or Rv2097c protein contained in the above kit can be in any form, including but not limited to direct protein samples, and the form of proteins immobilized on a certain carrier, such as a detection chip.

In addition to the Rv0389 protein, the Rv1408 protein and/or the Rv2097c protein, the kit at least includes kit instructions.

Judgment criteria for the results recorded in the instructions: using the combined detection of three proteins, if at least two of the anti-Rv0389 antibodies, anti-Rv1408 antibodies and/or anti-Rv2097c antibodies are positive in the test results of the sample to be tested, then It is judged that the sample to be tested is positive for active tuberculosis, otherwise it is tuberculosis latent infection.

Another aspect of the present invention is to provide a use of the above kit in the preparation of products for identifying, diagnosing and/or screening tuberculosis latent infection and/or active pulmonary tuberculosis.

The beneficial effects of the present invention are:

The invention can simultaneously detect the serum antibody response levels of three different Mycobacterium tuberculosis proteins, thereby better distinguishing and diagnosing tuberculosis latent infection patients and active tuberculosis patients, with higher sensitivity and better specificity.

Description of drawings

Figure 1 is a statistical chart of the expression levels of anti-Rv0389 antibody, anti-Rv1408 antibody and anti-Rv2097c antibody in the serum of latent tuberculosis infection patients (n=93) and active tuberculosis patients (n=92) detected by ELISA

Fig. 2 is the ROC curve diagram of joint detection of antibodies corresponding to three antigens;

sequence description

SEQ ID NO.1 is the amino acid sequence of the Rv0389 protein in the embodiment of the present invention;

SEQ ID NO.2 is the amino acid sequence of the Rv1408 protein in the embodiment of the present invention;

SEQ ID NO.3 is the amino acid sequence of the Rv2097c protein in the embodiment of the present invention.

Detailed ways

The invention discloses the use of a product for detecting the level of anti-Rv0389 antibody, anti-Rv1408 antibody and/or anti-Rv2097c antibody in a sample in the preparation of products for identifying, diagnosing and/or screening tuberculosis latent infection and/or active pulmonary tuberculosis. Those skilled in the art can refer to the content of this article to appropriately improve the process parameters to achieve. It needs to be pointed out that all similar replacements and modifications are obvious to those skilled in the art, and they are all considered to be included in the present invention, and relevant personnel can obviously make changes without departing from the content, spirit and scope of the present invention. Changes or appropriate changes and combinations are made to the content described herein to realize and apply the technology of the present invention.

In the present invention, unless otherwise specified, the scientific and technical terms used herein have the meanings commonly understood by those skilled in the art.

In order to enable those skilled in the art to better understand the technical solutions of the present invention, the present invention will be further described in detail below in conjunction with specific examples.

Example 1: Collection and processing of samples

The sample detected by the present invention is serum, and the collection method is to collect the blood of the subject on an empty stomach, use an ordinary biochemical test tube without anticoagulant, let it stand at room temperature until it is naturally coagulated, and then centrifuge at 3000 rpm for 10 minutes at room temperature to collect the serum fraction. Packed in cryopreservation tubes and stored in a -80°C low-temperature refrigerator 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 those who have no clinical complaint symptoms, TST test and T-SPOT test are positive, and chest X-ray films are negative; The active tuberculosis patient group refers to the confirmed tuberculosis patients with positive smear or culture, who have not started or just started systemic chemotherapy; both groups exclude diabetes, HIV infection and other autoimmune diseases.

The samples of the latent infection group used in the present invention come from the epidemiological investigation project of the Epidemiology Research Office of the Beijing Institute of Tuberculosis and Chest Tumors, and the samples of the active tuberculosis patient group come from inpatients in each ward of the Department of Tuberculosis, Beijing Chest Hospital Affiliated to Capital Medical University. This project has been approved by the Research Ethics Committee of Beijing Chest Hospital of Capital Medical University/Beijing Institute of Tuberculosis and Thoracic Tumors, and all research subjects have given informed consent.

Embodiment 2: Utilize ELISA experiment to the detection of sample

1. Establish various buffers, diluents, reaction solutions and stop solutions required for ELSIA experiments:

(1) Coating buffer (pH9.6, 0.05mol/L carbonate buffer):

Na ₂ CO ₃ 1.59g, NaHCO ₃ 2.93g, add double distilled water to 1000ml, adjust pH to 9.6;

(2) PBS solution with pH7.4:

NaCl 8.0g, KCl 0.2g, KH ₂ PO ₄ 0.2g, Na ₂ HPO ₄ 12H ₂ O 2.9g, add double distilled water to 1000ml, adjust pH to 7.4;

(3) PBST washing solution with pH7.4:

Add 0.5ml Tween-20 to 1000ml PBS solution, adjust the pH to 7.4;

(4) Blocking solution (pBS solution containing skimmed milk powder at pH 7.4):

Add 5g skimmed milk powder to 100ml PBS solution;

(5) Substrate buffer: 0.2M NaH ₂ PO ₄ (28.4g/L) 25.7ml,

0.1M citric acid (19.2g/L) 24.3ml, add 50ml of distilled water.

(6) TMB (tetramethylbenzidine) use solution: TMB (10mg/5ml absolute ethanol) 0.5ml substrate buffer (pH5.5) 10ml 0.75% H ₂ O ₂ 32μl;

(7) Stop solution (2M H ₂ SO ₄ ): 21.32ml H ₂ SO ₄ , 178.68ml H ₂ O.

(8) Secondary antibody: goat anti-human IgG labeled with horseradish peroxidase (HRP);

(9) 96-well ELISA plate; Nunc Incorporated (Theromo, Gebenhagen, Denmark)

2. Detection steps:

(1) Coating: Rv0389, Rv1408 and Rv2097c antigens were respectively dissolved in the coating buffer (5ug/ml); 100ul/well was added to a 96-well microtiter plate, and overnight at 4°C;

(2) Plate washing: PBST washing solution 300ul/well, 3min, wash 3 times;

(3) Blocking: blocking solution, 300ul/well, incubate at room temperature for 1h;

(4) Plate washing: washing solution 300ul/well, 3min, wash 3 times;

(5) Add serum: 1:400 diluted serum to be tested, 100ul/well, incubate at room temperature for 1h;

(6) Plate washing: wash solution 300ul/well, 3min, wash 5 times;

(7) Add enzyme-labeled secondary antibody: freshly diluted enzyme-labeled antibody (1:30000) 100ul/well, incubate at room temperature for 1h;

(8) Plate washing: wash solution 300ul/well, 3min, wash 5 times;

(9) Color development: freshly prepared TMB color development solution, 100ul/well, incubate at room temperature in the dark for 5-10min;

(10) Termination: stop solution 2M H ₂ SO ₄ , 50ul/well;

(11) Determination: Adjust the microplate reader to 450nm, measure the OD value of each well after zeroing the first blank PBS control well.

3. Result analysis and judgment criteria:

Use SPSS22.0 software to draw the ROC curve, and according to the coordinate values, the specificity and sensitivity corresponding to different critical points can be obtained, and then the Youden index can be calculated (Youden index = specificity + sensitivity - 1), and three antigens can be obtained The critical point OD value of Rv0389 is 0.291, Rv1408 is 0.283, Rv2097c is 0.308;

The results are shown in Figure 1, and the results indicated that all three proteins have the function of distinguishing tuberculosis latent infection from active tuberculosis.

judgement standard:

Using the combined detection of three proteins, if at least two of the anti-Rv0389 antibody, anti-Rv1408 antibody and/or anti-Rv2097c antibody are positive in the test results of the test sample, it is determined that the test sample is positive for active tuberculosis , otherwise latent tuberculosis infection.

Example 3: Specificity and sensitivity of the method for differential diagnosis of active tuberculosis patients and latent infections using ELISA

Adopt clinical diagnosis as 92 parts of serum of patients with active pulmonary tuberculosis, 93 parts of serum of latent infection; Utilize the ELISA method among the embodiment 1 and 2 to detect, judge whether the test subject is active pulmonary tuberculosis positive by the judgment standard of embodiment 2 , the results are shown in Figure 2, the specificity of the results analysis was 86.0%, and the sensitivity was 71.7%. In Figure 2, the abscissa 1-specificity represents the false positive rate, and the ordinate sensitivity represents the true positive rate.

The above results prove that the present invention detects the serum antibody response levels of three different Mycobacterium tuberculosis proteins, can better differentiate and diagnose latent tuberculosis infection patients and active tuberculosis patients, and has higher sensitivity and better specificity.

The above is only a preferred embodiment of the present invention, it should be pointed out that, for those of ordinary skill in the art, without departing from the principle of the present invention, some improvements and modifications can also be made, and these improvements and modifications can also be made. It should be regarded as the protection scope of the present invention.

sequence listing

<110> Beijing Chest Hospital Affiliated to Capital Medical University

<120> Use of Mycobacterium tuberculosis protein in the preparation of products for diagnosing tuberculosis latent infection and/or active pulmonary 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 70 75 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

290 295 300

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 (4)

1. Use of a product for detecting levels of an anti-Rv 0389 antibody, an anti-Rv 1408 antibody and an anti-Rv 2097c antibody in a sample for the preparation of a product for the identification, diagnosis and/or screening of tuberculosis latent infection;

the sample is serum.

2. A use according to claim 1, wherein the products for detecting levels of anti-Rv 0389 antibodies, anti-Rv 1408 antibodies and anti-Rv 2097c antibodies in a sample comprise Rv0389 protein, rv1408 protein and Rv2097c protein.

3. The use according to claim 2, wherein the Rv0389 protein is shown in SEQ ID No. 1; the Rv1408 protein is shown in SEQ ID NO. 2; the Rv2097c protein is shown as SEQ ID NO. 3.

4. The use according to any one of claims 1 to 3, wherein the method of detection comprises an enzyme-linked immunosorbent assay, an agglutination assay, a precipitation assay, an E-garland assay, a small phagocytosis assay, an immunofluorescence assay, a colloidal gold immunochromatography assay, a spot gold immunodiafiltration assay and/or an electrochemiluminescence assay.

Images