Polypeptide combination and application thereof

Abstract

The invention relates to the technical field of biology, in particular to a polypeptide combination and application thereof. The invention provides a human giant cell IgM epitope polypeptide. The invention also provides a method for coupling the epitope polypeptide with horse radish peroxidase. The invention also provides a detection method of the epitope polypeptide coupled horseradish peroxidase applied to the human giant cell IgM detection kit. The method has the advantages that the two epitope polypeptides are identified for the first time to detect positive serum samples of human cytomegalovirus IgM, the human cytomegalovirus IgM detection kit adopting the polypeptides as specific antigens has excellent sensitivity and specificity, and the standard is easy to control.

Description

Technical Field

[0001] The present invention relates to the field of biotechnology, and in particular to a polypeptide combination and application thereof. Background Art

[0002] Human cytomegalovirus (HCMV) belongs to the genus cytomegalovirus of the family Herpesviridae, also known as human beta herpesvirus type 5. The viral genome is a double-stranded DNA molecule of 200 kb in length. The virus is an enveloped virus with a particle diameter of 200 nm. From the outside to the inside, it consists of an envelope, a vesicle, and a nucleocapsid. The nucleocapsid is T16 icosahedral symmetry and contains 162 repeated capsomeres.

[0003] Human cytomegalovirus is widely transmitted in the human population, with an infection rate of 60-70% among adults in developed countries and over 90% in developing countries. HCMV infection is mainly caused by direct contact with body fluids, including tears, saliva, genital secretions and blood. HCMV is an immunosuppressive virus that causes almost no symptoms in healthy people, but for people with weakened immune function, such as HIV-infected people, organ transplant recipients or newborns, HCMV infection can be life-threatening.

[0004] Human cytomegalovirus can cause intrauterine infection of the fetus through vertical transmission from mother to child. HCMV infection is an important viral cause of neonatal birth defects. Intrauterine infection of the fetus with HCMV is the main cause of deafness, intellectual disability and learning disabilities in newborns. Statistics show that the probability of intrauterine infection of HCMV in newborns is about 0.5%, of which 5% may have multiple disorders and cytomegalovirus inclusion disease with nonspecific signs similar to rubella. Therefore, the diagnosis of HCMV infection in pregnant women in the early pregnancy is of great significance in the field of eugenics.

[0005] Since HCMV produces almost no symptoms in the normal population and can recur intermittently in an asymptomatic form, HCMV infection is difficult to diagnose through clinical symptoms. People infected with HCMV will produce antibodies against the virus, especially IgM antibodies. Serological tests, especially human cytomegalovirus IgM tests, have important clinical significance for indicating early and recurrent human cytomegalovirus infection.

[0006] The main antigens detected by human giant cell serological testing are viral proteins such as G52 and pp150. Among them, pp150 is the most abundant protein in the HCMV virus particle envelope. IgM antibodies produced against pp150 appear in the early stage of HCMV infection. G52 is a non-structural protein produced during the replication of the virus in cells. Due to the high HCMV IgG positive rate in adults, human giant cell IgM testing using G52 or pp150 as specific antigens is susceptible to various interferences such as rheumatoid factors and target-specific IgG antibodies, resulting in a certain false positive rate. Summary of the invention

[0007] In view of this, the polypeptide combination and application provided by the present invention have excellent sensitivity and specificity, and the standards are easy to control.

[0008] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0009] The present invention provides the use of antigenic epitopes in any of the following items:

[0010] (a) preparing a human cytomegalovirus IgM detection reagent or kit;

[0011] (b) Preparation of human cytomegalovirus diagnostic reagents or kits;

[0012] The antigenic epitope includes antigenic epitope 1 and / or antigenic epitope 2;

[0013] The antigenic epitope 1 has:

[0014] (1) the amino acid sequence shown in SEQ ID NO: 1; or

[0015] (2) An amino acid sequence obtained by substituting, deleting or adding one or more residues of the amino acid sequence shown in (1), and having the same or similar function as (1); or

[0016] (3) an amino acid sequence having at least 90% homology to the amino acid sequence shown in (1) or (2);

[0017] The antigenic epitope 2 has:

[0018] (4) the amino acid sequence shown in SEQ ID NO: 2; or

[0019] (5) An amino acid sequence obtained by substituting, deleting or adding one or more residues of the amino acid sequence shown in (4), and having the same or similar function as (4); or

[0020] (6) an amino acid sequence having at least 90% homology to the amino acid sequence shown in (4) or (5);

[0021] The plurality is 2 to 5.

[0022] The present invention also provides a polypeptide combination, including polypeptide 1 and polypeptide 2;

[0023] The polypeptide 1 has:

[0024] (1) the amino acid sequence shown in SEQ ID NO: 1; or

[0025] (2) An amino acid sequence obtained by substituting, deleting or adding one or more residues of the amino acid sequence shown in (1), and having the same or similar function as (1); or

[0026] (3) an amino acid sequence having at least 80% homology to the amino acid sequence shown in (1) or (2);

[0027] The polypeptide 2 has:

[0028] (4) the amino acid sequence shown in SEQ ID NO: 2; or

[0029] (5) An amino acid sequence obtained by substituting, deleting or adding one or more residues of the amino acid sequence shown in (4), and having the same or similar function as (4); or

[0030] (6) an amino acid sequence having at least 80% homology to the amino acid sequence shown in (4) or (5);

[0031] The plurality is 2 to 5.

[0032] The present invention also provides a fusion polypeptide, comprising the above polypeptide combination and a connecting arm;

[0033] The connecting arm comprises a connecting arm 1 and a connecting arm 2;

[0034] The connecting arm 1 has:

[0035] (7) the amino acid sequence shown in SEQ ID NO: 3; or

[0036] (8) An amino acid sequence obtained by substituting, deleting or adding one or more residues of the amino acid sequence shown in (7), and having the same or similar function as (7); or

[0037] (9) an amino acid sequence having at least 80% homology to the amino acid sequence shown in (7) or (8);

[0038] The connecting arm 2 has:

[0039] (10), the amino acid sequence shown in SEQ ID NO: 4; or

[0040] (11) An amino acid sequence obtained by substituting, deleting or adding one or more residues of the amino acid sequence shown in (10), and having the same or similar function as (10); or

[0041] (12) an amino acid sequence having at least 80% homology to the amino acid sequence shown in (10) or (11);

[0042] The plurality is 2 to 3.

[0043] In some specific embodiments of the present invention, the above-mentioned fusion polypeptide comprises:

[0044] The nitrogen-terminal residue of the linker 1 is modified with pimeloyl-biotin; and / or

[0045] The lysine residues in the linker 2 are modified with biotin.

[0046] In some specific embodiments of the present invention, the above-mentioned fusion polypeptide includes fusion polypeptide 1 and fusion polypeptide 2;

[0047] The fusion polypeptide 1 is formed by the fusion of the polypeptide 1 and the connecting arm;

[0048] The fusion polypeptide 2 is formed by the fusion of the polypeptide 2 and the connecting arm.

[0049] In some specific embodiments of the present invention, the above-mentioned fusion polypeptide comprises:

[0050] The fusion polypeptide 1 comprises:

[0051] (i) formed by connecting the nitrogen terminus of the polypeptide 1 to the carbon terminus of the linker 1; and / or

[0052] (ii), formed by connecting the carbon terminus of the polypeptide 1 to the nitrogen terminus of the connecting arm 2;

[0053] and / or

[0054] The fusion polypeptide 2 comprises:

[0055] (iii), formed by connecting the nitrogen terminus of the polypeptide 2 to the carbon terminus of the connecting arm 1; and / or

[0056] (iv) The carbon terminus of the polypeptide 2 is connected to the nitrogen terminus of the connecting arm 2.

[0057] In some specific embodiments of the present invention, the above-mentioned fusion polypeptide has:

[0058] (13) an amino acid sequence as shown in any of SEQ ID NOs: 5 to 8; or

[0059] (14) An amino acid sequence obtained by substituting, deleting or adding one or more residues of the amino acid sequence shown in (13), and having the same or similar function as (13); or

[0060] (15) An amino acid sequence having at least 80% homology to the amino acid sequence shown in (13) or (14);

[0061] The plurality is 2 to 5.

[0062] The present invention also provides the use of the above fusion polypeptide in the following aspects:

[0063] (a) preparing a human cytomegalovirus IgM detection reagent or kit;

[0064] (b) Preparation of human cytomegalovirus diagnostic reagents or kits.

[0065] The present invention also provides a reagent or a kit, comprising the above polypeptide combination or the above fusion polypeptide, and acceptable auxiliary materials or adjuvants.

[0066] The present invention also provides a detection method, comprising:

[0067] (I) using the above polypeptide combination to detect the sample to be tested to obtain the test result; or

[0068] (II), using the above fusion polypeptide to detect the sample to be tested to obtain the test result; or

[0069] (III) Using the above reagents or kits to test the sample to obtain the test results.

[0070] The polypeptide combination and application of the present invention have the following effects:

[0071] 1. Two short peptides on the human cytomegalovirus structural protein pp150 were identified for the first time. The antigenic epitopes corresponding to these two short peptides are the main epitopes of human cytomegalovirus IgM antibodies.

[0072] 2. Through a specific labeling method, the two polypeptide epitopes bound to the horseradish peroxidase luminescence system can be fully exposed.

[0073] 3. Human giant cell IgM capture kit was assembled by short peptides labeled with horseradish peroxidase, which can specifically identify human giant cell IgM positive serum.

[0074] 4. The results of comparative testing using the HCMV IgM detection kit of DiaSorin as a standard showed that the specificity of the kit of the present invention was 100% and the sensitivity was 98%, which was better than the HCMV IgM kit of a domestic company (98% and 96.7%, respectively). Although the sensitivity was slightly lower than that of the HCMV IgM detection kit of DiaSorin (100%), the specificity was higher than that of the HCMV IgM detection kit of DiaSorin and the HCMV IgM kit of a domestic company (96.7% and 94%, respectively). BRIEF DESCRIPTION OF THE DRAWINGS

[0075] Figure 1 The chemical structure of pimeloyl-biotin is shown. DETAILED DESCRIPTION

[0076] The present invention discloses polypeptide combinations and their applications. Those skilled in the art can refer to the content of this article and appropriately improve the process parameters to achieve them. It should be particularly noted that all similar substitutions and modifications are obvious to those skilled in the art and are considered to be included in the present invention. The methods and applications of the present invention have been described through preferred embodiments, and relevant personnel can obviously modify or appropriately change and combine the methods and applications described herein without departing from the content, spirit and scope of the present invention to implement and apply the technology of the present invention.

[0077] The purpose of the present invention is to provide a separated polypeptide in view of the shortcomings of the existing human HCMV IgM detection technology;

[0078] The second object of the present invention is to provide another polypeptide;

[0079] The third object of the present invention is to provide a polypeptide-based linker;

[0080] The fourth object of the present invention is to provide a method for labeling the above-mentioned polypeptide linker;

[0081] The fifth object of the present invention is to provide a method for labeling horseradish peroxidase with a polypeptide;

[0082] The sixth object of the present invention is to provide a kit for detecting human giant cell IgM antibodies;

[0083] To achieve the above first object, the technical solution adopted by the present invention is: an isolated polypeptide, wherein the polypeptide sequence is selected from any one of the amino acid sequences shown in SEQ ID NO: 1~2.

[0084] To achieve the above second purpose, the technical solution adopted by the present invention is: a polypeptide, the polypeptide sequence:

[0085] a) a tandem arrangement of any one of the amino acid sequences shown in SEQ ID NO: 1 to 2; or

[0086] b) a combination of any two of the amino acid sequences shown in SEQ ID NO: 1 to 2,

[0087] The polypeptide is used to prepare a diagnostic reagent for diagnosing human HCMV infection. In the polypeptide, the tandem polypeptides, i.e., the polypeptides whose sequences are shown in SEQ ID NO: 1-2, are randomly selected in order and in the number of copies of each polypeptide.

[0088] To achieve the third object, the technical solution adopted by the present invention is: an isolated polypeptide, wherein the polypeptide sequence is selected from any one of the amino acid sequences shown in SEQ ID NO: 3~4.

[0089] To achieve the fourth purpose, the technical solution adopted by the present invention is: the modification method of the polypeptide connecting arm is:

[0090] a) the nitrogen terminus of SEQ ID NO: 3 is modified with pimeloyl-biotin; or

[0091] b) the lysine in SEQ ID NO: 4 is a biotin-modified lysine;

[0092] in:

[0093] a) the amino acid in SEQ ID NO: 3 is fused to the nitrogen terminus of the polypeptides shown in SEQ ID NO: 1-2; or

[0094] b) the amino acid in SEQ ID NO: 4 is fused to the carbon terminus of the polypeptide shown in SEQ ID NO: 1-2;

[0095] To achieve the fifth purpose, the technical solution adopted by the present invention is: the biotinylated modified polypeptide is co-incubated with streptavidin-coupled horseradish peroxidase (SA-HRP), and then the unbound polypeptide is removed by dialysis to prepare a polypeptide enzyme label.

[0096] To achieve the sixth objective, the technical solution adopted by the present invention is: a capture kit for detecting human cytomegalovirus IgM, the kit comprising any of the polypeptides or molecules described above.

[0097] The human cytomegalovirus antigen polypeptide proposed by the present invention has an amino acid sequence as shown in SEQ ID NO: 1-2.

[0098] The polypeptide connecting arm proposed in the present invention has an amino acid sequence as shown in SEQ ID NO: 3-4.

[0099] The modification method of the polypeptide coupling arm proposed by the present invention is:

[0100] a) the nitrogen terminus of SEQ ID NO: 3 is modified with pimeloyl-biotin; or

[0101] b) the lysine in SEQ ID NO: 4 is a biotin-modified lysine;

[0102] The fusion method of the modified polypeptide coupling arm and the antigen polypeptide proposed by the present invention is:

[0103] a) the amino acid in SEQ ID NO: 3 is fused to the nitrogen terminus of the polypeptides shown in SEQ ID NO: 1-2; or

[0104] b) the amino acid in SEQ ID NO: 4 is fused to the carbon terminus of the polypeptide shown in SEQ ID NO: 1-2;

[0105] The human cytomegalovirus modified antigen peptide sequence proposed by the present invention has an amino acid sequence as shown in SEQ ID NO: 5-8.

[0106] The modified antigen polypeptide proposed in the invention was synthesized by Shanghai Sangon Biotechnology Co., Ltd. with a purity of more than 99% and was verified to be qualified by mass spectrometry.

[0107] The modified antigen polypeptide and horseradish peroxidase coupling method proposed in the invention is: polypeptide co-incubation streptavidin coupled HRP.

[0108] The streptavidin-HRP conjugated to the present invention is provided by Zhengzhou Imino Biotechnology Co., Ltd.

[0109] The sequence information involved in the present invention is as follows:

[0110] SEQ ID NO: 1 is TAPAPAPTPTFAGTQT;

[0111] SEQ ID NO: 2 is AKTPSDAVQNILQKIEKIKNTEE;

[0112] SEQ ID NO: 3 is GGGSGG;

[0113] SEQ ID NO: 4 is GGGSGGK;

[0114] SEQ ID NO: 5 is GGGSGGTAPAPAPTPTFAGTQT;

[0115] SEQ ID NO: 6 is TAPAPAPTPTFAGTQTGGGSGGK;

[0116] SEQ ID NO: 7 is GGGSGGAKTPSDAVQNILQKIEKIKNTEE;

[0117] SEQ ID NO: 8 is AKTPSDAVQNILQKIEKIKNTEEGGGSGGK;

[0118] Unless otherwise specified, the raw materials, reagents, consumables and instruments involved in the present invention are all common commercial products and can be purchased from the market.

[0119] The present invention will be further described below in conjunction with embodiments:

[0120] Example 1

[0121] Preparation of human giant cell IgM antibody detection kit:

[0122] (1) Synthesis of biotin-modified human cytomegalovirus antigen peptide

[0123] The sequence of the human cytomegalovirus biotin-modified antigen polypeptide is shown in SEQ ID NO: 5-8, which was commissioned to Shanghai Sangon Biotechnology Co., Ltd. for synthesis, with a purity of greater than 98%, and was freeze-dried for storage.

[0124] (2) Biotin-modified human cytomegalovirus antigen peptide coupled to horseradish peroxidase

[0125] Take 1 mg of human cytomegalovirus antigen peptide and dissolve it in 2 ml ultrapure water to make a 0.5 mg / ml peptide solution. Add 2 ml of the peptide solution to 40 ml of streptavidin-coupled HRP solution and mix well. Rotate and mix well at 37°C for 2 hours. The reactant is dialyzed into PBS buffer containing 50% glycerol at a dialysis ratio of 1:200. The solution is changed once during the dialysis to obtain a human cytomegalovirus antigen peptide-horseradish peroxidase conjugate.

[0126] (3) A human cytomegalovirus IgM antibody detection kit

[0127] A human cytomegalovirus IgM antibody detection kit comprises magnetic particles coupled with anti-μ secondary antibodies, sample diluent, enzyme-labeled antigen stock solution, enzyme-labeled antigen diluent, washing solution, substrate A solution, substrate B solution, negative control substance, positive control substance and instructions.

[0128] In this embodiment:

[0129] The magnetic particles coupled to the anti-μ secondary antibody were prepared by Zhengzhou Antu Biotechnology Co., Ltd.

[0130] The enzyme-labeled antigen stock solution is the above-mentioned biotin-modified human cytomegalovirus antigen polypeptide mixed with streptavidin-coupled HRP solution.

[0131] Preparation of negative control: Collect more than 6 normal human sera that are negative for cytomegalovirus IgM antibody, HBsAg, HIV (1+2) antibody, TP antibody, and HCV antibody detected by ELISA method, filter and sterilize, aliquot, and store at low temperature.

[0132] Preparation of positive control: Collect more than 6 normal human sera that are positive for cytomegalovirus IgM antibody and negative for HBsAg, HIV (1+2) antibody, TP antibody, and HCV antibody detected by ELISA method, filter and sterilize, aliquot, and store at low temperature.

[0133] Sample diluent (10 mM PBS buffer, 0.6% BSA, 0.2% casein, 0.1% ProClin300, 20% calf serum, pH 7.2)

[0134] Substrate Solution A and Substrate Solution B are prepared by Zhengzhou Antu Biological Co., Ltd.

[0135] Enzyme-labeled antigen diluent (10 mM PBS buffer, 0.6% BSA, 0.2% casein, 0.1% ProClin300, 2% BSA, 0.1% Triton X-100, pH 7.2)

[0136] Specifically, dilute the enzyme-labeled antigen stock solution 10,000 times with the enzyme-labeled antigen diluent.

[0137] The detection instrument uses the A2000 plus chemiluminescence analyzer of Zhengzhou Antu Biological Co., Ltd.

[0138] The detection program uses the human cytomegalovirus IgM detection program provided with the A2000 plus chemiluminescence analyzer of Zhengzhou Antu Biological Co., Ltd.

[0139] Example 2

[0140] Method for using the human cytomegalovirus IgM antibody detection kit

[0141] (1) Equilibrate the human cytomegalovirus IgM antibody detection kit prepared in Example 1 at room temperature for 15-30 minutes.

[0142] (2) Place the test sample in the Zhengzhou Antu sample rack and perform sample detection using the human cytomegalovirus IgM detection program.

[0143] (3) Cutoff value = 2.1 * (NC average value). If the RLU value > Cutoff value, the sample is determined to be a positive sample; if the RLU value < Cutoff value, the sample is determined to be a negative sample.

[0144] Example 3

[0145] The human cytomegalovirus IgM antibody detection kit prepared in Example 1 was used to detect 90 normal human blood samples and 10 clinically confirmed cytomegalovirus acute infection samples. The detection procedure was strictly carried out according to the method provided in Example 2. The results are as follows:

[0146] Table 1: Clinical trial evaluation results of the kit of the present invention

[0147]

[0148] The data described in Table 1 show that the specificity and sensitivity of the kit of the present invention are excellent, and the positive predictive value and negative predictive value of the kit of the present invention are both 100%.

[0149] The data in Table 1 indicate that the human cytomegalovirus antigen polypeptide epitopes provided by the present invention can effectively detect clinical human cytomegalovirus IgM positive samples.

[0150] The human cytomegalovirus antigen polypeptide site provided by the invention is simple and effective, and has good kit performance.

[0151] Example 4: Comparison of the above kit with the HCMV IgM kit (indirect method) of DiaSorin and the HCMV IgM kit (capture method) of a domestic company:

[0152] HCMV IgM kit (capture method) from a domestic company:

[0153] 1. The kit consists of an anti-human IgMμ chain antibody-coated enzyme-labeled reaction plate, positive control serum, negative control serum, specimen diluent, washing solution, color developing solution, and stop solution. The specific components are:

[0154] (1) HCMV IgM positive and negative sera;

[0155] (2) Specimen diluent: 0.01M pH 7.4 phosphate buffered saline (PBS) containing 10% calf serum and plasma chromogen

[0156] (3) Washing solution: phosphate buffered saline containing 0.05% Tween 20 (PBST);

[0157] (4) Enzyme-labeled antigen: recombinant protein pp150 labeled with horseradish peroxidase;

[0158] (5) Color developing solution: TMB-H2O2 urea solution;

[0159] (6) Stop solution: 2 mol / L H2SO4 solution.

[0160] 2. Usage:

[0161] (1) Add the serum to be tested at a dilution of 1:100 to the coated ELISA reaction plate, 100 μl / well, incubate at 37°C for 1 h, and wash the plate 3 times;

[0162] (2) Add enzyme-labeled antigen and dilute HRP-pp150 to the working concentration, 100 μl / well, incubate at 37°C for 30 min.

[0163] (3) Color development: After washing, add 100 μl of TMB-H2O2 system substrate to each well and develop color at 37°C or room temperature in the dark for 15 min;

[0164] (4) Add 50 μl of 2 mol / L H2SO4 to each well to terminate the reaction;

[0165] (5) Result judgment: adjust the blank well to zero and read the absorbance at a wavelength of 450 nm, i.e., the OD450 value.

[0166] Comparison method of this embodiment:

[0167] (1) Take clinically confirmed HCMV IgM positive and negative sera, test them according to the detection methods specified in the instructions of the three kits, and make positive and negative judgments according to the positive and negative judgment criteria in the kit instructions.

[0168] (2) Calculate the specificity and sensitivity of the kit of the present invention, the HCMV IgM kit of DiaSorin Company, and the HCMV IgM kit of a domestic company.

[0169] Table 2 is a comparison of the kit of the present invention with the HCMV IgM detection kit of DiaSorin and the HCMV IgM kit of a domestic company.

[0170] Table 2

[0171]

[0172] The results of comparative testing using the HCMV IgM detection kit of DiaSorin as a standard showed that the specificity of the kit of the present invention was 100% and the sensitivity was 98%, which was better than the HCMV IgM kit of a domestic company (98% and 96.7%, respectively). Although the sensitivity was slightly lower than that of the HCMV IgM detection kit of DiaSorin (100%), the specificity was higher than that of the HCMV IgM detection kit of DiaSorin and the HCMV IgM kit of a domestic company (96.7% and 94%, respectively).

[0173] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. Application of antigenic epitopes in any of the following: (a) preparing a human cytomegalovirus IgM detection reagent or kit; (b) Preparation of human cytomegalovirus diagnostic reagents or kits; The antigenic epitope includes antigenic epitope 1 and / or antigenic epitope 2; The antigenic epitope 1 has: (1) the amino acid sequence shown in SEQ ID NO: 1; or (2) An amino acid sequence obtained by substituting, deleting or adding one or more residues of the amino acid sequence shown in (1), and having the same or similar function as (1); or (3) an amino acid sequence having at least 90% homology to the amino acid sequence shown in (1) or (2); The antigenic epitope 2 has: (4) the amino acid sequence shown in SEQ ID NO: 2; or (5) An amino acid sequence obtained by substituting, deleting or adding one or more residues of the amino acid sequence shown in (4), and having the same or similar function as (4); or (6) an amino acid sequence having at least 90% homology to the amino acid sequence shown in (4) or (5); The plurality is 2 to 5.

2. A polypeptide combination, characterized in that: Includes peptide 1 and peptide 2; The polypeptide 1 has: (1) the amino acid sequence shown in SEQ ID NO: 1; or (2) An amino acid sequence obtained by substituting, deleting or adding one or more residues of the amino acid sequence shown in (1), and having the same or similar function as (1); or (3) an amino acid sequence having at least 80% homology to the amino acid sequence shown in (1) or (2); The polypeptide 2 has: (4) the amino acid sequence shown in SEQ ID NO: 2; or (5) An amino acid sequence obtained by substituting, deleting or adding one or more residues of the amino acid sequence shown in (4), and having the same or similar function as (4); or (6) an amino acid sequence having at least 80% homology to the amino acid sequence shown in (4) or (5); The plurality is 2 to 5.

3. A fusion polypeptide, characterized in that Comprising the polypeptide combination as claimed in claim 2 and a connecting arm; The connecting arm comprises a connecting arm 1 and a connecting arm 2; The connecting arm 1 has: (7) the amino acid sequence shown in SEQ ID NO: 3; or (8) An amino acid sequence obtained by substituting, deleting or adding one or more residues of the amino acid sequence shown in (7), and having the same or similar function as (7); or (9) an amino acid sequence having at least 80% homology to the amino acid sequence shown in (7) or (8); The connecting arm 2 has: (10), the amino acid sequence shown in SEQ ID NO: 4; or (11) An amino acid sequence obtained by substituting, deleting or adding one or more residues of the amino acid sequence shown in (10), and having the same or similar function as (10); or (12) an amino acid sequence having at least 80% homology to the amino acid sequence shown in (10) or (11); The plurality is 2 to 3.

4. The fusion polypeptide according to claim 3, characterized in that include: The nitrogen-terminal residue of the linker 1 is modified with pimeloyl-biotin; and / or The lysine residues in the linker 2 are modified with biotin.

5. The fusion polypeptide according to claim 3 or 4, characterized in that: Including fusion polypeptide 1 and fusion polypeptide 2; The fusion polypeptide 1 is formed by the fusion of the polypeptide 1 and the connecting arm; The fusion polypeptide 2 is formed by the fusion of the polypeptide 2 and the connecting arm.

6. The fusion polypeptide according to claim 5, characterized in that include: The fusion polypeptide 1 comprises: (i) formed by connecting the nitrogen terminus of the polypeptide 1 to the carbon terminus of the linker 1; and / or (ii), formed by connecting the carbon terminus of the polypeptide 1 to the nitrogen terminus of the connecting arm 2; and / or The fusion polypeptide 2 comprises: (iii), formed by connecting the nitrogen terminus of the polypeptide 2 to the carbon terminus of the connecting arm 1; and / or (iv) The carbon terminus of the polypeptide 2 is connected to the nitrogen terminus of the connecting arm 2.

7. The fusion polypeptide according to any one of claims 3 to 6, characterized in that have: (13) an amino acid sequence as shown in any of SEQ ID NOs: 5 to 8; or (14) An amino acid sequence obtained by substituting, deleting or adding one or more residues of the amino acid sequence shown in (13), and having the same or similar function as (13); or (15) An amino acid sequence having at least 80% homology to the amino acid sequence shown in (13) or (14); The plurality is 2 to 5.

8. Use of the polypeptide combination according to claim 2 or the fusion polypeptide according to any one of claims 3 to 7 in the following aspects: (a) preparing a human cytomegalovirus IgM detection reagent or kit; (b) Preparation of human cytomegalovirus diagnostic reagents or kits.

9. A reagent or a kit, characterized in that It comprises the polypeptide combination as claimed in claim 2 or the fusion polypeptide as claimed in any one of claims 3 to 7, and acceptable excipients or adjuvants.

10. A detection method, characterized in that: include: (I) using the polypeptide combination as claimed in claim 2 to detect the sample to be tested and obtain the test result; or (II), using the fusion polypeptide according to any one of claims 3 to 7 to detect the sample to be tested, and obtaining a test result; or (III) Detecting the sample to be tested using the reagent or kit as described in claim 9 to obtain a test result.

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