Test strip for immunochromatography comprising hepatitis d antigen protein and diagnostic kit comprising same

Abstract

The present invention relates to a diagnostic kit for hepatitis D, which detects an antibody against hepatitis D virus (HDV) in a sample to rapidly and accurately detect the presence or absence of HDV infection and, more specifically, to a diagnostic kit for hepatitis D, comprising a recombinant HDV antigen.

Description

Test strip for immunochromatography containing hepatitis D antigen protein and diagnostic kit containing the same

[0001] The present invention relates to a hepatitis D diagnostic kit capable of detecting infection with HDV, and more specifically, to a hepatitis D diagnostic kit containing an antigen of HDV.

[0002] Hepatitis D virus (HDV), also known as delta virus, is a small RNA virus that can cause a severe form of hepatitis in humans. It differs from other hepatitis viruses in that it requires the hepatitis B virus (HBV) to replicate to cause infection. HDV can infect individuals through co-infection with HBV or through superinfection in individuals already chronically infected with HBV.

[0003]

[0004] If hepatitis D is suspected, a blood test is performed to confirm the diagnosis. This test detects antibodies produced by the body's immune system in response to the hepatitis D virus. HDV infection is diagnosed by performing both total anti-HDV and IgG anti-HDV tests simultaneously. IgM anti-HDV is known to appear not only in the acute phase but also in the chronic phase, limiting its diagnostic utility. It can result in false negatives in immunosuppressed patients and false positives in healthy individuals, as anti-HDV remains after recovery from acute hepatitis D.

[0005]

[0006] Meanwhile, immunochromatography (ICA) has recently become increasingly popular due to its advantages of being able to obtain simple and quick results, enabling the measurement of various types of substances in various fields, including antigens, antibodies, hormones, and drugs.

[0007] Against this backdrop, it is essential to select a more accurate and sensitive method for diagnosing hepatitis D.

[0008] Accordingly, the inventors of the present invention sought to provide a test strip for immunochromatography capable of quickly and easily diagnosing HDV infection even with a small amount of sample, and a diagnostic kit including the test strip.

[0009] To achieve the above purpose,

[0010] In one aspect, the present invention provides a composition for detecting hepatitis D virus antibodies comprising a recombinant hepatitis D virus (HDV).

[0011] In another aspect, an immunochromatographic strip comprising the composition for detecting the hepatitis D virus antibody is provided.

[0012] In one embodiment, the strip comprises:

[0013] (a) a sample pad into which the sample is absorbed;

[0014] (b) a conjugation pad comprising a gold conjugate;

[0015] (c) a membrane including a test line on which a recombinant hepatitis D virus antigen protein is immobilized and a control line on which a secondary antibody is immobilized;

[0016] (d) an absorption pad into which the remaining sample is absorbed, and

[0017] (e) a strip for immunochromatography, including a support;

[0018] It may include, and the composition for detecting the hepatitis B virus antibody may be included in the binding pad.

[0019] In one embodiment, the gold particles in the gold conjugate may be colloidal gold particles and may have a particle size of 20 to 50 nm.

[0020] In another aspect of the present invention, a kit for diagnosing hepatitis D virus or detecting hepatitis D virus antibodies comprising the above-described immunochromatographic strip is provided.

[0021] In another aspect of the present invention, in order to provide information necessary for the diagnosis of hepatitis D virus, an analysis method is provided, which uses the composition for detecting hepatitis D virus to perform a method selected from the group consisting of an immunochromatography method, an immunodot assay method, an analysis method using a Luminex assay system, and a protein microarray method.

[0022] As described above, using the hepatitis D diagnostic kit of the present invention, HDV infection can be diagnosed within a short period of time. Furthermore, the diagnostic kit of the present invention can diagnose HDV virus infection simply and economically, using a subject's blood, urine, feces, or other bodily fluids, without the need for expensive samples, equipment, or skilled techniques.

[0023]

[0024] Figure 1 is a drawing showing the immunochromatographic working principle of the HDV antibody rapid measurement kit according to the present invention.

[0025] Figure 2 is a diagram showing the information of the recombinant HDV antigen protein, the epitope site, and the SDS PAGE results of the purified recombinant HDV antigen.

[0026] Figure 3 is a drawing showing the configuration of an immunochromatography immunoassay strip according to the present invention.

[0027] Figure 4 is a photograph showing the rapid test results of positive and negative controls for the HDV antibody detection assay test.

[0028] Figures 5a-e are photographs showing the results of analyzing a sample of group A using the HDV antibody rapid measurement kit according to the present invention.

[0029] Figures 6a-b are photographs showing the results of analyzing a sample from group B using the HDV antibody rapid measurement kit according to the present invention.

[0030] Figure 7 is a photograph showing the results of analyzing a sample of group C using the HDV antibody rapid measurement kit according to the present invention.

[0031] The present invention relates to a composition for detecting hepatitis D virus antibodies comprising a hepatitis D virus (HDV) antigen protein. Specifically, the HDV antigen protein may be a recombinant HDV antigen protein.

[0032] In one embodiment, the recombinant HDV antigen protein may be at least one selected from the group consisting of the protein of SEQ ID NO: 1 and proteins immunologically equivalent thereto. The "immunologically equivalent protein" is one in which a corresponding amino acid residue of a protein or enzyme is changed without altering the overall structure and function of the polypeptide, such as by replacing a certain amino acid with an amino acid having similar properties (e.g., polarity, hydrogen bonding potential, acidity, basicity, hydrophobicity, aromaticity, etc.). The "immunologically equivalent protein" also includes a polypeptide that has at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% amino acid homology as measured by the BLAST or FASTA algorithm, and has the same or substantially similar properties or functions as the native protein or parent protein to which it is compared. In one embodiment, the production of the recombinant antigen protein of the HDV may be accomplished by a method of PCR amplifying a DNA sequence encoding the antigen, cloning it into an expression vector, expressing the antigen protein, and isolating and purifying the expressed protein.

[0033] In a more specific embodiment, the recombinant HDV antigen protein comprises a T cell epitope represented by a sequence selected from the group consisting of SEQ ID NOs: 2-4 and a B cell epitope represented by SEQ ID NO: 5.

[0034]

[0035] [T cell epitope]

[0036] Sequence number 2: GGREEILEQWVAGRKKLEEL

[0037] Sequence number 3: LKKIEDENPWLGNIKGILGK

[0038] Sequence number 4: AKRARTDQMEADSGPGKRPL

[0039]

[0040] [B cell epitope]

[0041] Sequence number 5: RTGEGLDIRGNQGFP

[0042]

[0043] Specifically, in the present invention, a recombinant HDV antigen protein forms an antigen-antibody complex with an antibody against HDV contained in a sample, specifically a biological sample, and the formed antigen-antibody complex is detected.

[0044] The term “biological sample” used in the present invention includes tissues, cells, whole blood, serum, plasma, saliva, cerebrospinal fluid, urine, etc. The expression level of hepatitis D virus can be measured by reacting these biological samples with or without manipulation with the recombinant HDV antigen protein of the present invention or an antigen protein containing the same epitope. In the present invention, the “antigen-antibody complex” refers to a combination of the hepatitis D virus and the HDV antibody contained in the sample, which are reacted to confirm the expression of HDV in the biological sample. The formation of the antigen-antibody complex can be detected by a method selected from the group consisting of a colorimetric method, an electrochemical method, a fluorimetric method, a luminometry method, a particle counting method, a visual assessment method, and a scintillation counting method. However, it is not necessarily limited to these and various applications and applications are possible.

[0045] In the present invention, various markers are used to detect antigen-antibody complexes. Specific examples include enzymes, fluorescent substances, ligands, luminescent substances, microparticles, and radioactive isotopes. However, the present invention is not limited to these. Enzymes used as detection labels include acetylcholinesterase, alkaline phosphatase, β-D galactosidase, horseradish peroxidase (HRP), β-latamase, etc.; fluorescent substances include fluorescein, Eu 3+, Eu 3+ chelate, or cryptate, etc.; ligands include biotin derivatives, etc.; luminescent substances include acridinium ester, isoluminol derivatives, etc.; microparticles include colloidal gold particles, colored latex, etc.; and radioactive isotopes include 57 Co, 3 H, 125 I, 125 I-Bonton Hunter reagent, etc. The particle size of the gold particles may be preferably 20 to 50 nm, more preferably 30 to 40 nm, and most preferably 35 nm.

[0046] In another aspect of the present invention, the present invention provides an analysis method for performing a method selected from the group consisting of an immunochromatography method, an immunodot assay method, an analysis method using a Luminex assay system, and a protein microarray method, using the composition for detecting HDV antibodies, in order to provide information necessary for the diagnosis of HDV.

[0047] In one embodiment, the analytical method for measuring HDV antibodies performed to provide information necessary for the diagnosis of HDV in the present invention is preferably a method using immunochromatography or a Luminex assay system.

[0048] Immunochromatographic analysis is a method recently used for point-of-care testing in the field of diagnostic immunology. When a test substance is present in a test sample, a fusion of a specific binding substance capable of specifically binding to the test substance and a label having labeling properties forms a complex with the test substance on a stationary phase on which a specific binding substance capable of specifically binding to the test substance is immobilized, and the presence of the test substance in the test sample can be confirmed by detecting the label bound to the stationary phase. Therefore, in the case of the analysis method used to provide information for HDV diagnosis according to the present invention, an antibody specific for HDV in a biological sample (anti-HDV antibody) is the test substance, and when this test substance is applied to the sample path and begins to migrate, it reacts with Protein A bound to the label and continuously develops in the form of an antibody-protein A complex. As it migrates, the recombinant HDV antigen protein immobilized on the reaction membrane and the antibody-protein A complex react once more to form a sandwich-type complex. Since the recombinant HDV antigen protein is immobilized on the reaction membrane, when the antigen-antibody reaction continues, the complex accumulates on the immobilized surface of the capture antibody. Since the protein is transparent to the naked eye, the presence or absence of the complex and the relative amount can be determined by the presence or absence of the hepatitis D virus through the fused label, preferably a chromogenic substance capable of binding to the antibody, and more preferably, a gold particle containing the chromogenic substance. Preferably, in the present invention, an immunochromatographic assay can be performed using an immunochromatographic strip containing a recombinant HDV antigen or a kit containing the same.

[0049] Accordingly, as another aspect of the present invention, the present invention provides an immunochromatographic strip comprising the composition for detecting hepatitis D virus.

[0050] The method for manufacturing an immunochromatography strip can be performed according to a conventional strip manufacturing method. Preferably, the immunochromatography strip according to the present invention may include (a) a sample pad on which a sample is absorbed, (b) a conjugation pad containing a chromogenic substance capable of binding to an antibody, (c) a reaction membrane including a test line on which a recombinant hepatitis D virus (recombinant HDV) antigen protein is immobilized and a control line on which a secondary antibody is immobilized, (d) an absorption pad on which the remaining sample is absorbed, and (e) a support. The composition for detecting the hepatitis D virus antibody may be included in the test line in a form in which the recombinant hepatitis D virus antigen protein is immobilized.

[0051] The sample pad is used to apply a sample, i.e., the above-mentioned biological sample, by a drop or the like and absorb it into the strip, and the absorbed sample passes through the binding pad by capillary action and moves along the reaction membrane. The binding pad includes a chromogenic substance capable of binding to an antibody as a label, for example, a colloidal gold conjugate including a chromogenic substance, and more preferably, a gold-protein A conjugate in which colloidal gold including a chromogenic substance is conjugated to protein A, and a gold-Chicken IgY antibody conjugate in which colloidal gold including a chromogenic substance is conjugated to Chicken IgY antibody.

[0052] The reaction membrane is preferably a nitrocellulose (NC) membrane and / or a glass fiber (GF) membrane.

[0053] Additionally, an absorbent pad is positioned at the opposite end of the membrane and absorbs the biological sample that has moved along the membrane by capillary action, and the support is preferably made of plastic to support the components of this strip.

[0054]

[0055] In another aspect, the present invention provides a kit for diagnosing hepatitis D virus or detecting HDV antibodies, comprising the immunochromatographic strip. The immunochromatographic strip of the present invention is preferably in the form of a cassette or stick. The immunochromatographic strip and the diagnostic kit comprising the same are very useful for analysis to provide information for the diagnosis of HDV, and have the advantage of being able to perform analysis very quickly and accurately even with low concentrations of HDV antibodies.

[0056]

[0057] Hereinafter, the present invention will be described in more detail through examples and test examples. However, the following examples and test examples are intended to illustrate the present invention and are not intended to limit the scope of the present invention.

[0058] Example

[0059]

[0060] Example 1. Preparation of recombinant HDV antigen

[0061] By comparing the genotypes of several human delta viruses and analyzing epitopes, we selected s-HDAg, which exhibits high sensitivity and specificity. Furthermore, we modified the nucleic acid sequence through codon usage analysis to suit the E. coli expression system, resulting in a 195-amino acid sequence, represented by SEQ ID NO: 1.

[0062]

[0063] A recombinant protein is produced using the BL21(DE3) / pGEX 4T-1 / HDV clone, which is transformed into BL21(DE3) after ligating a DNA fragment containing the sequence of human Delta virus into the pGEX 4T-1 vector. In a 1 L flask, add 250 mL of LB medium, 250 μL of ampicillin (100 mg / ml), and 5 mL of the BL21(DE3) / pGEX 4T-1 / HDV culture that was grown overnight. Place the flask in a shaking incubator, and set the temperature to 37°C and the shaking speed to 200 rpm. When the OD600 value reaches 0.5, store in the refrigerator for 30 minutes. Dispense 125 μL of 1 M IPTG and place it in the shaking incubator. Set the shaking incubator to 24℃ and the shaking speed to 200rpm and culture for 6 hours. Transfer the culture medium to a 250ml centrifuge tube and centrifuge at 10,000xg, 4℃, and 5 minutes. After centrifugation, discard the supernatant and resuspend the pellet in 30ml of 1xPBS solution. Disrupt the cells in the resuspended solution by sonicating the sonicator at 50% output, 10 seconds on, 30 seconds off, and repeating 10 times. Transfer the disrupted solution to a 50ml centrifuge tube and centrifuge at 20,000xg, 4℃, and 20 minutes. After centrifugation, carefully transfer the supernatant to a new tube to prevent the pellet from being carried away and filter using a 0.45um syringe filter. The filtered solution is separated and purified using a GSTrap column. The separated and purified protein is buffer-exchanged to 20 mM carbonate buffer using Amicon® Ultra-15 Centrifugal Filter Units (30 kDa).After buffer exchange, HDV recombinant protein is quantified using the BCA method. Electrophoresis is performed on a 12% SDS-PAGE gel to confirm that the HDV recombinant protein is at least 90% pure. The HDV recombinant protein, whose concentration and purity have been confirmed, is aliquoted and stored frozen.

[0064]

[0065] Example 2. Preparation of a kit containing an immunostrip (HDV Ag Rapid Kit)

[0066] An immune strip containing the antigen protein of the expressed recombinant HDV and a kit containing the same were manufactured to diagnose hepatitis D. The immune strip is composed of a sample pad; a binding pad; a nitrocellulose (hereinafter also referred to as “NC”) membrane; and an absorbent pad, wherein the sample pad, binding pad, membrane, and absorbent pad are fixed on a solid support so as to overlap each other, thereby forming a single immune strip as a whole (Fig. 1).

[0067]

[0068] A 2.54 cm long nitrocellulose membrane was prepared by bonding it to a backbone with an adhesive on the plastic backing surface. Here, Recombinant HDV was dispensed at regular intervals on the lower part of the nitrocellulose (NC) membrane and goat-derived anti-chicken IgY was dispensed at regular intervals on the upper part using a line dispenser (LA C-Production-03) that dispenses a fixed amount of solution, and then physically immobilized by drying in a drying chamber for more than 3 hours. At this time, the secondary antibody at the control line position in the upper part can be applied without any other effect on the system even if other polyclonal antibodies such as rabbit-derived anti-mouse IgG are used.

[0069] A solution prepared by conjugating protein A and colloidal gold was soaked in a glass fiber membrane at a certain concentration, dried in a drying chamber for more than 3 hours, and cut into a certain size to prepare a gold pad. At this time, the concentration of the sucrose component was maintained at 5% or more so that the deposited protein A-gold could easily dissolve in the moving solution and escape from the pad. In addition, casein or BSA, which is one of the inactive proteins, was added and deposited, and then dried together to prepare.

[0070] The sample pad, through which the solution is introduced into the analysis system, was prepared using cellulose fiber or glass fiber. When the solution is introduced into the analysis system, the components present in the sample must migrate without physical or chemical reactions occurring before binding to the protein-gold on the gold pad or antibodies immobilized on the NC membrane.

[0071] The absorption pad located at the top of the analysis system is made of cellulose fiber (or cotton fiber can be used) and is made of a cellulose membrane with a certain thickness or more so that it can absorb the solution supplied from the bottom so that the solution participating in the reaction can maintain a continuous flow. This is configured so that the solution participating in the reaction can continuously progress from the bottom of the system and move through the signal reading section.

[0072]

[0073] Example 3. Confirmation of signal generation pattern of inspection line

[0074] In order to conduct experiments to identify the components contributing to the movement of the solution and the signal in lateral flow, we designed a strip-type system using recombinant antigens and antibodies against HDV at the signal measurement site and attempted to identify the patterns contributing to signal generation.

[0075] In the lateral flow system, the test line consists of recombinant, gold-conjugated protein A.

[0076] The recombinant was immobilized by precipitating it on the NC membrane at a concentration of 0.4–1.2 mg / mL and used as the signal measurement site. The protein A used in the reaction was prepared by combining it with colloidal gold so that the amount was 0.47 mg / mL. The antibody used in the experiment was a monoclonal HDV antibody produced by our company, and the antibody of the control line was a goat-derived anti-chicken IgY antibody (1.0 mg / mL) and the signal generation pattern was confirmed.

[0077] When the system is configured in this way, when a signal generation reaction occurs, first, when the solution is supplied from the bottom of the system, the components present at each location can mix to cause a reaction and move. It was confirmed that the signal generation source protein A used in the experiment and the HDV antibody contained in the sample reacted and moved, and reacted with the recombinant HDV in the test line, which is the signal generation part for the antibody, to generate a signal. In the case of HDV negativity, since there is no HDV antibody in the sample, the protein A+gold particle that is not bound to the antibody moves. It was confirmed that another signal generation part, goat anti-chicken IgY antibody in the control line, and the signal generation polyclonal antibody chicken IgY reacted and generated a signal. It was confirmed that a difference in the signal occurred depending on the change in the concentration of the monoclonal HDV antibody used.

[0078]

[0079] Example 4. Serum sample testing using the HDV antibody rapid test kit according to the present invention.

[0080] To determine whether the kit according to the present invention can rapidly detect HDV antibodies, detection of HDV antibodies was confirmed using 200 serum samples.

[0081] A total of 200 serum samples were obtained from the Mongolian Liver Center (ONOM Foundation, Mongolia). All samples were pre-tested within the clinical service framework using commercially available anti-HCV and HBsAg ELISAs and a self-developed HDV-RNA quantitative PCR kit. Serum samples were divided into the following groups:

[0082]

[0083] Group A: 121 HDV-RNA positive serum samples.

[0084] Group B: 50 HDV-RNA negative, HBsAg and HBV-DNA positive serum samples.

[0085] Group C: 30 HBsAg and anti-HCV negative serum samples

[0086]

[0087] A 10 μl sample of serum was collected using a micropipette. The micropipette was held at a 90° angle to the test device, and the collected sample was added to the test well. The buffer bottle was held at a 90° angle to the test device, taking care not to touch the test well, and three drops of buffer were added to the test well. The results were checked after 15 minutes.

[0088]

[0089] First, the rapid test results using the HDV antibody detection kit according to the present invention for group A samples (see Table 1) showed that all samples were positive.

[0090] The rapid test results using the HDV antibody detection kit according to the present invention for group B samples (see Table 2) showed that most samples were negative, with one sample (B19) showing a positive reaction. The B19 serum sample that showed a positive reaction was retested using the commercially available Wantai anti-HDV IgG kit and confirmed to be indeed positive, and was therefore moved to group A (positive).

[0091] The rapid test results using the HDV antibody detection kit according to the present invention for group C samples (see Table 3) showed that most samples were negative, and one sample (C2) tested positive. The C2 serum sample that tested positive was retested using the commercially available Wantai anti-HDV IgG kit and was confirmed to be positive, so it was moved to group A (positive). Accordingly, the samples were divided into group A: 122, group B: 49, and group C: 29, and statistical analysis was performed.

[0092] According to the statistical analysis results, there were 122 true positives, 78 true negatives, and 0 false positives and false negatives.

[0093] Table 1. LUCA HDV Ab rapid test result for Group A samples

[0094]

[0095] Table 2. LUCA HDV Ab rapid test results for Group B samples SampleIDStandard resultHDV-RNAIU / mlStandard resultHBV-DNAIU / mlStandard resultHBsAg IU / mlRapid test resultB10132259.08PositiveNegativeB2012400-NegativeB3002.62PositiveNegativeB406181159.04 PositiveNegativeB50-47111.63 PositiveNegativeB60246014248.52 PositiveNegativeB701520-NegativeB8067601181.87PositiveNegativeB907950556.23 PositiveNegativeB10013-NegativeB110463NegativeB120-167.01 PositiveNegativeB130-321.97 PositiveNegativeB140-0.44 PositiveNegativeB15002.62 PositiveNegativeB160-6611.82 PositiveNegativeB1701770000230.08PositiveNegativeB180542096.83PositiveNegativeB190746-PositiveB200-4547.54PositiveNegativeB2101016772.56PositiveNegativeB22078200-NegativeB230-8295.47PositiveNegativeB240--NegativeB25010-NegativeB260-1093.04PositiveNegativeB2701980002893.07PositiveNegativeB2801980002893.07PositiveNegativeB29050304909.79 PositiveNegativeB30050304909.79 PositiveNegativeB3102380256.87 PositiveNegativeB3202380256.87 PositiveNegativeB330-321.97 PositiveNegativeB3403370-NegativeB350-204.73PositiveNegativeB3605540-NegativeB370-99306.34PositiveNegativeB380-3457.84 PositiveNegativeB390340-NegativeB400181000000-NegativeB41051-NegativeB420792-NegativeB4302460014248.52PositiveNegativeB4401520-NegativeB45067601181.87PositiveNegativeB460255-NegativeB47050200-NegativeB48076-NegativeB490-1975.7NegativePositiveB500463-Negative.

[0096]

[0097] Table 3. LUCA HDV Ab rapid test results for Group C samplesSample IDStandard result HBsAg ELISAStandard resultAnti-HCV ELISARapid test resultC1NegativeNegativeNegativeC2NegativeNegativePositiveC3NegativeNegativeNegativeC4NegativeNegativeNegativeC5NegativeNegativeNegativeC6NegativeNegativeNegativeC7NegativeNegativeNegativeC8NegativeNegativeNegativeC9NegativeNegativeNegativeC10NegativeNegativeNegativeC11NegativeNegativeNegativeC12NegativeNegativeNegativeC13NegativeNegativeNegativeC14NegativeNegativeNegativeC15NegativeNegativeNegativeC16NegativeNegativeNegativeC17NegativeNegativeNegativeC18NegativeNegativeNegativeC19NegativeNegativeNegativeC20NegativeNegativeNegativeC21NegativeNegativeNegativeC22NegativeNegativeNegativeC23NegativeNegativeNegativeC24NegativeNegativeNegativeC25NegativeNegativeNegativeC26NegativeNegativeNegativeC27NegativeNegativeNegativeC28NegativeNegativeNegativeC29NegativeNegativeNegativeC30NegativeNegativeNegative

[0098]

[0099] 표준 결과양성음성총합본 발명에 따른 HDV 진단 키트의 항체 시험 결과양성1220122음성07878총합12278200

[0100] With these results, sensitivity, specificity, and accuracy were calculated according to the formula below.

[0101]

[0102] (TP: true positive; TN: true negative; FP: false positive; FN: false negative)

[0103]

[0104] As a result of the test, among the samples from groups B and C that were negative in the pre-test of each serum sample before testing with the kit according to the present invention, two samples tested positive using the kit according to the present invention, indicating that these were false negative samples. These two samples were further confirmed as HDV-RNA positive using the commercial Wantai HDV Ab IgG kit. Thus, it was confirmed that the kit according to the present invention exhibits excellent accuracy.

[0105]

[0106] In addition, as a result of calculating sensitivity, specificity, and accuracy as described above, it was found that when using the kit according to the present invention, sensitivity, specificity, and accuracy were all 100%.

Claims

1. A composition for detecting hepatitis D virus antibody comprising a hepatitis D virus (HDV) antigen protein.

2. A composition for detecting hepatitis D virus antibodies, characterized in that in the first paragraph, the antigen protein is a protein represented by an amino acid sequence of sequence number 1 or an amino acid sequence having 70% or more, 80% or more, or 90% or more homology with the amino acid sequence.

3. An immunochromatographic strip comprising a composition for detecting hepatitis D virus antibodies according to Article 1. 4.(a) Sample pad where the sample is absorbed; (b) a conjugation pad containing a chromogenic substance to which an antibody can bind; (c) a membrane including a test line on which a recombinant hepatitis D virus antigen protein is immobilized and a control line on which a secondary antibody is immobilized; (d) an absorption pad into which the remaining sample is absorbed, and (e) A strip for immunochromatography comprising a support.

5. In paragraph 4, An immunochromatographic strip, wherein the chromogenic substance to which the above antibody can bind is a gold conjugate.

6. In paragraph 5, An immunochromatographic strip, wherein the gold conjugate is selected from the group consisting of a gold-protein A conjugate in which colloidal gold containing a chromogenic substance is conjugated to protein A, a gold-Chicken IgY antibody conjugate in which colloidal gold containing a chromogenic substance is conjugated to Chicken IgY antibody, and a mixture thereof.

7. In paragraph 4, A strip for immunochromatography, wherein the above reaction membrane is a nitrocellulose membrane or a glass fiber membrane.

8. A kit for detecting hepatitis D virus antibodies comprising an immunochromatographic strip according to Article 4.

9. A diagnostic kit for hepatitis D virus comprising an immunochromatographic strip according to Article 4.

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