Application of TFF1 and TFF3 in the early diagnosis of lung cancer bone metastasis

Abstract

The present invention belongs to the field of biomedicine technology, discloses the application of TFF1 and TFF3 in the early diagnosis of lung cancer bone metastasis, and specifically discloses the application of a substance for detecting biomarkers in the preparation of a product for diagnosis or early screening of lung cancer bone metastasis; the biomarkers include TFF1 and / or TFF3. The present invention discloses for the first time the relationship between the expression of secretory proteins TFF1 and TFF3 and specific bone metastasis of lung cancer, and verifies that secretory proteins TFF1 and TFF3 are specifically highly expressed in serum specimens of patients with lung cancer bone metastasis and lung cancer bone metastasis cells, suggesting that secretory proteins TFF1 and TFF3 can be used as a new diagnostic marker for lung cancer bone metastasis, which can significantly distinguish lung cancer patients with bone metastasis from lung cancer patients with metastasis to other organs and healthy people.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine technology, and specifically relates to the application of TFF1 and TFF3 in the early diagnosis of lung cancer bone metastasis. Background Art

[0002] The incidence and mortality of lung cancer rank first among all tumors in the world. For a long time, the survival rate of lung cancer patients has been extremely low, especially for patients with mid-late and advanced lung cancer, with a five-year survival rate of only 9% and 1% respectively. And in actual clinical diagnosis, up to 60% to 80% of patients are already in the advanced stage at the beginning of diagnosis, with a poor prognosis. Metastasis is the cause of death in more than 90% of cancer patients, among which bone is the most common and most severe metastatic organ of advanced lung cancer. The low early diagnosis rate is considered to be the main reason for the high mortality rate and poor prognosis of patients with lung cancer bone metastasis.

[0003] Bone metastasis usually has no symptoms in the early stage, and its symptoms are related to the location and number of tumor metastases. When patients seek medical treatment with typical symptoms of bone metastasis such as bone pain and pathological fractures, they have usually developed into larger metastatic lesions. In clinical practice, bone imaging is often used to determine whether tumor patients have bone metastasis, such as radionuclide bone scanning, PET, PET-CT and MRI, but these methods have defects such as high cost and delayed diagnosis, and have certain limitations in sensitivity, specificity and operability.

[0004] In recent years, research on biomarkers for the diagnosis of bone metastasis has made some progress. Researchers have found that compared with prostate cancer patients without bone metastasis, patients with bone metastasis have significantly increased serum levels of ProGRP. Through prospective studies, researchers found that the appearance of osteocalcin-positive circulating tumor cells means that patients are more likely to develop bone metastasis. In patients with advanced lung cancer, the expression levels of proteins such as CXCR4, BSP, OPN, and BMP4 in tumor cells are positively correlated with the risk of bone metastasis. However, the above studies are currently in the preclinical research stage, and there are currently no bone metastasis-specific tumor markers for clinical use, especially lung cancer. Therefore, exploring the molecular mechanism of lung cancer bone metastasis and finding new, highly sensitive and easy-to-detect early diagnostic markers for bone metastasis are of great clinical value and are very urgent. Summary of the invention

[0005] The purpose of the first aspect of the present invention is to provide the use of a substance for detecting biomarkers in the preparation of a product for the diagnosis or early screening of lung cancer bone metastasis.

[0006] The second aspect of the present invention aims to provide the use of TFF1 and / or TFF3 as biomarkers in the preparation of products for early diagnosis of lung cancer bone metastasis.

[0007] The third aspect of the present invention aims to provide the use of inhibitory agents of TFF1 and / or TFF3 in the preparation of drugs for preventing, inhibiting and / or treating bone metastasis of lung cancer.

[0008] The fourth aspect of the present invention aims to provide the use of TFF1 and / or TFF3 as targets in screening drugs for treating or preventing bone metastasis of lung cancer.

[0009] The fifth aspect of the present invention aims to provide a marker combination.

[0010] The sixth aspect of the present invention aims to provide a product.

[0011] In order to achieve the above object, the technical solution adopted by the present invention is:

[0012] The first aspect of the present invention provides the use of a substance for detecting a biomarker in the preparation of a product for diagnosing or early screening of lung cancer bone metastasis; the biomarker includes TFF1 and / or TFF3.

[0013] Preferably, the substance includes a substance for quantitatively detecting a biomarker.

[0014] Preferably, the substance for detecting a biomarker comprises a substance for detecting a biomarker at a gene level and / or a protein level.

[0015] Preferably, the substance for detecting biomarkers is a substance used in one or more detection techniques or methods selected from the following group: enzyme-linked immunosorbent assay, immunofluorescence, radioimmunoassay, immunocoprecipitation, immunoblotting, high performance liquid chromatography, capillary gel electrophoresis, near infrared spectroscopy, mass spectrometry, immunochemiluminescence, colloidal gold immunoassay, fluorescent immunochromatography, surface plasmon resonance, immuno-PCR or biotin-avidin technology.

[0016] Preferably, the substance for detecting biomarkers is selected from: a substance specific to TFF1 and / or TFF3, a probe specific to TFF1 and / or TFF3, a gene chip, a PCR primer, and the like.

[0017] Preferably, the substance specific to TFF1 and / or TFF3 is any one of (b1) to (b3):

[0018] (b1) an antibody that specifically binds to TFF1 and / or TFF3;

[0019] (b2) a ligand protein or polypeptide that specifically binds to TFF1 and / or TFF3;

[0020] (b3) Non-protein compounds that specifically recognize TFF1 and / or TFF3.

[0021] Preferably, the antibody comprises at least one of a polyclonal antibody, a monoclonal antibody, a single-chain antibody, a functional antibody fragment, an antibody Fab region, a nanobody, a chimeric antibody, and a multispecific antibody.

[0022] Preferably, the product comprises at least one of a reagent, a test kit, a test paper, and a chip.

[0023] Preferably, the sample of the subject of the product may be selected from a body fluid sample, a tumor sample and a cell sample isolated from the subject.

[0024] Preferably, the body fluids include but are not limited to blood, plasma, serum, lymph, cerebrospinal fluid, synovial fluid, urine, saliva, mucus, etc.; the forms of the tumor samples include but are not limited to living tissue, paraffin-embedded tissue, frozen tissue, etc.; the cell samples can be selected from cells that can be isolated from a subject, such as peripheral blood mononuclear cells, T lymphocytes, B lymphocytes, circulating tumor cells, etc.

[0025] Preferably, when TFF1 and / or TFF3 of the subject to be tested is significantly increased relative to a reference level, it is diagnosed as lung cancer bone metastasis; the reference level is the level of subjects of the same age group who do not suffer from lung cancer bone metastasis.

[0026] Preferably, the subjects of the same age group are healthy people and lung cancer patients without bone metastasis.

[0027] The term "no bone metastasis" in the medical field refers to the absence of bone metastasis. Generally speaking, the collected tumor samples are analyzed, and if the tumor sample has no bone metastasis, it is classified as a tumor sample without bone metastasis. Those skilled in the art should know that a tumor sample without bone metastasis may be a tumor sample without any metastasis, or may be a tumor sample with other non-bone metastases (such as liver, brain, and lung).

[0028] Preferably, the lung cancer includes non-small cell lung cancer and small cell lung cancer.

[0029] Preferably, the non-small cell lung cancer includes squamous cell carcinoma (SCC), adenocarcinoma, and large cell carcinoma.

[0030] Preferably, the expression of TFF1 and / or TFF3 is upregulated in patients with lung cancer bone metastasis.

[0031] The second aspect of the present invention provides the use of TFF1 and / or TFF3 as a biomarker in the preparation of a product for the early diagnosis of lung cancer bone metastasis.

[0032] Preferably, the product includes at least one of a reagent, a test kit, a test paper, and a chip.

[0033] The third aspect of the present invention provides the use of inhibitory agents of TFF1 and / or TFF3 in the preparation of drugs for preventing, inhibiting and / or treating bone metastasis of lung cancer.

[0034] Preferably, the inhibitory agent of TFF1 and / or TFF3 includes a substance that can fully or partially inhibit the expression of TFF1 and / or TFF3 genes, and / or a substance that inhibits the function of TFF1 and / or TFF3 proteins.

[0035] Preferably, the inhibitory agent of TFF1 and / or TFF3 is selected from: proteins, oligonucleotides, oligonucleotide expression vectors, and small molecule compounds.

[0036] Preferably, the expression vector comprises a eukaryotic cell expression vector, and the eukaryotic cell expression vector comprises a plasmid expression vector or a viral expression vector.

[0037] A fourth aspect of the present invention provides the use of TFF1 and / or TFF3 as targets in screening drugs for treating or preventing bone metastasis of lung cancer.

[0038] According to a fifth aspect of the present invention, a marker combination is provided, wherein the marker combination comprises TFF1 and TFF3.

[0039] Preferably, the marker combination consists of TFF1 and TFF3.

[0040] The sixth aspect of the present invention provides a product comprising a substance for detecting the marker combination of the fifth aspect of the present invention.

[0041] The beneficial effects of the present invention are:

[0042] The present invention discloses for the first time the relationship between the expression of secretory proteins TFF1 and TFF3 and specific bone metastasis of lung cancer, and verifies that secretory proteins TFF1 and TFF3 are specifically and highly expressed in serum specimens of patients with lung cancer bone metastasis and in lung cancer bone metastasis cells, suggesting that secretory proteins TFF1 and TFF3 can be used as a new diagnostic marker for lung cancer bone metastasis, which can significantly distinguish lung cancer patients with bone metastasis from lung cancer patients with metastasis to other organs and healthy people.

[0043] In clinical applications, ELISA can be used to detect the levels of TFF1 and TFF3 in patient blood samples to determine the risk of bone metastasis in patients so as to implement earlier intervention treatment. Compared with traditional bone metastasis diagnosis methods such as tissue biopsy or imaging screening, ELISA detection found that the expression levels of TFF1 and TFF3 in the serum of lung cancer bone patients were significantly increased, indicating that it indicates the possibility of bone metastasis in patients, so that relevant treatment can be carried out more promptly and bring better prognosis to patients; from the perspective of operation, it is more convenient and easy, the results are more sensitive, and the experimental cost is lower; from the patient's perspective, the psychological and economic burdens of serum ELISA non-invasive detection are much smaller. BRIEF DESCRIPTION OF THE DRAWINGS

[0044] Figure 1 is the expression of TFF1 and TFF3 in lung cancer bone metastasis cells and serum of patients with lung cancer bone metastasis; wherein, A is the expression of TFF1 and TFF3 in lung cancer bone metastasis cells, B is the expression of TFF1 in the serum of patients with lung cancer bone metastasis, C is the expression of TFF3 in the serum of patients with lung cancer bone metastasis; D is the ROC curve of TFF1 for the diagnosis of lung cancer bone metastasis, E is the ROC curve of TFF3 for the diagnosis of lung cancer bone metastasis, and F is the ROC curve of TFF1 and TFF3 combined for the diagnosis of lung cancer bone metastasis.

[0045] Figure 2 This is the result of an in vivo experiment in mice to detect the effect of TFF1 and TFF3 expression on bone metastasis of lung cancer. DETAILED DESCRIPTION

[0046] The following will be combined with the embodiments of the present application to clearly and completely describe the technical solutions in the embodiments of the present application. Obviously, the described embodiments are only part of the embodiments of the present application, not all of the embodiments. Based on the embodiments in the present application, all other embodiments obtained by those skilled in the art without creative work are within the scope of protection of the present application.

[0047] Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the art. Unless otherwise specified, the reagents and materials used in the following examples are commercially available. Experimental methods without specific conditions are usually carried out under conventional conditions or conditions recommended by the manufacturer.

[0048] Example 1 Expression of TFF1 and TFF3 in lung cancer bone metastasis cells and serum of lung cancer bone metastasis patients

[0049] The inventor team screened in mice and extracted PC9 bone metastasis (PC9-BM) cells and PC9 lung metastasis (PC9-LM) cells, and found that DKK3 was significantly highly expressed in PC9-BM cells, suggesting that the expression of secretory protein DKK3 is related to lung cancer bone metastasis and can be used as a blood molecular diagnostic marker for early lung cancer bone metastasis. The inventor used enzyme-linked immunosorbent assay (ELISA) to detect the expression of TFF1 and TFF3 in the supernatant of lung cancer-specific organ metastasis subcell lines and the serum of lung cancer bone metastasis patients, and verified the specific secretion of TFF1 and TFF3 in bone metastasis cells and the serum of lung cancer bone metastasis patients, which specifically includes the following steps:

[0050] 1. Collect cell supernatant / extract patient serum

[0051] Collect cell supernatant: 2×10 6 Density inoculated PC9 parental cells (PC9-PR), PC9-BM cells, and PC9-LM cells in 100×20 mm culture dishes, respectively; cultured at 37°C and 5% CO2 for 24 hours, discarded the culture medium, washed twice with PBS, and discarded PBS; added 8 mL of serum-free culture medium (Opti-MEM, Gibco, Catalog: #31985070) and continued to culture; after 48 hours, collected the cell supernatant and filtered with a 0.45 μm filter to remove cell debris in the cell supernatant; collected the cell filtrate and immediately carried out subsequent experiments or stored at -40°C for later use.

[0052] Extract patient serum (separate serum from freshly collected whole blood): collect whole blood from 20 healthy subjects, 33 patients with early lung cancer without metastasis, and 45 patients with lung cancer with bone metastasis (from the Cancer Prevention and Treatment Center of Sun Yat-sen University) in anticoagulant tubes (containing EDTA), and let it stand at 4°C overnight; after standing, stratification can be seen, the lower layer is plasma, and the upper layer is serum; aspirate the upper clear yellow serum layer into a centrifuge tube, and centrifuge at 4°C, 2500rpm for 30 minutes; discard the precipitate, aspirate the upper serum component, and obtain the patient serum, which can be used immediately for subsequent experiments or stored at -80°C for later use.

[0053] 2.ELISA detection of TFF1 and TFF3 expression in cell supernatant and serum samples

[0054] The expression levels of TFF1 and TFF3 in cell supernatants and serum samples were detected using R&D Duoset series ELISA kits (R&D, Catalog: #DY5237, #DY4407). The specific operation is as follows:

[0055] (1) After the capture antibody (R&D, PART: #843815, #843133) was diluted to a working concentration (2 mg / mL) using PBS solution, 100 μL was added to each well of an ELISA 96-well plate and incubated at room temperature (20-30°C) overnight;

[0056] (2) Discard the capture antibody in the ELISA 96-well plate and wash each well three times with 400 μL of ELISA wash buffer. After the last wash, turn the ELISA 96-well plate upside down on absorbent paper and tap the plate gently to completely remove the residual ELISA wash buffer in the well.

[0057] (3) Add 300 mL of Reagent Diluent to each well and incubate at room temperature for 1 hour for blocking treatment;

[0058] (4) Discard the Reagent Diluent in the wells and wash each well three times with 400 μL ELISA wash buffer each time; after the last wash, turn the ELISA 96-well plate upside down on absorbent paper and tap the plate gently to completely remove the residual ELISA wash buffer in the wells;

[0059] (5) According to the instructions, dilute the TFF1 protein standard (R&D, PART: #843817) to 15.6 pg / mL, 31.3 pg / mL, 62.5 pg / mL, 125 pg / mL, 250 pg / mL, 500 pg / mL, and 1000 pg / mL, and dilute the TFF3 protein standard (R&D, PART: #843135) to 7.8 pg / mL, 15.6 pg / mL, 31.3 pg / mL, 62.5 pg / mL, 125 pg / mL, 250 pg / mL, and 500 pg / mL. Add the diluted protein standards and the samples to be tested to a 96-well plate, add 100 μL to each well, and incubate at room temperature for 2 hours;

[0060] (6) Discard the samples in the wells and wash each well three times with 400 μL ELISA wash buffer. After the last wash, place the ELISA 96-well plate upside down on absorbent paper and tap the plate gently to completely remove the residual ELISA wash buffer in the wells.

[0061] (7) Add 100 μL of TFF1 detection antibody (R&D, PART: #843816, concentration of 100 ng / mL) or TFF3 detection antibody (R&D, PART: #843134, concentration of 100 ng / mL) to each well and incubate at room temperature for 2 hours;

[0062] (8) Discard the liquid in the wells and wash each well three times with 400 μL ELISA wash buffer. After the last wash, turn the ELISA 96-well plate upside down on absorbent paper and tap the plate gently to completely remove the residual ELISA wash buffer in the wells.

[0063] (9) Add 100 μL of Streptavidin-HRP (200×) to each well and incubate at room temperature for 20 min in the dark;

[0064] (10) Discard the liquid in the wells and wash each well three times with 400 μL ELISA wash buffer. After the last wash, place the ELISA 96-well plate upside down on absorbent paper and tap the plate gently to completely remove the residual ELISA wash buffer in the wells.

[0065] (11) Add 100 μL of substrate solution to each well for color development. After adding the substrate solution, incubate at room temperature in the dark for 20 minutes.

[0066] (12) Add 50 μL of stop solution to each well to stop the reaction. After adding the stop solution, tap the plate to mix thoroughly.

[0067] (13) Immediately after termination, use a multifunctional microplate reader to measure the absorbance at 450 nm. 450nm The regression curve was calculated and the protein concentration of the sample was calculated.

[0068] The results are as follows Figure 1 As shown, TFF1 and TFF3 are specifically upregulated in lung cancer bone metastasis cells ( Figure 1 In serum samples of patients with lung cancer bone metastasis, TFF1 and TFF3 were specifically upregulated ( Figure 1 B-C), and was statistically significant (p<0.0001).

[0069] In order to further evaluate the value of secretory proteins TFF1 and TFF3 in the early diagnosis of lung cancer bone metastasis, the inventors constructed ROC curves for secretory proteins TFF1 and TFF3 to predict lung cancer bone metastasis in lung cancer patients with reference to the clinical pathological diagnosis results. Figure 1As shown in D, the AUC of secretory protein TFF1 in distinguishing bone metastasis from non-bone metastasis (including healthy and early non-metastasis) is 0.8897, 95% CI (0.8247-0.9548), the AUC in distinguishing healthy people from early non-bone metastasis is 0.6258, 95% CI (0.4636-0.7879), and the AUC in distinguishing early non-bone metastasis from bone metastasis is 0.9017, 95% CI (0.8374-0.966); Figure 1 As shown in E, the AUC of secretory protein TFF3 in distinguishing bone metastasis from non-bone metastasis (including healthy and early non-metastasis) was 0.9825, 95% CI (0.8247-0.9548), the AUC in distinguishing healthy people from early non-bone metastasis was 0.534, 95% CI (0.3751-0.6929), and the AUC in distinguishing early non-bone metastasis from bone metastasis was 0.9728, 95% CI (0.9373-1.008); when secretory protein TFF1 and TFF3 were used together, the AUC for distinguishing early non-bone metastasis from bone metastasis lung cancer patients was 0.987, 95% CI (0.966-1.000), and the AUC for distinguishing non-bone metastasis (including healthy and early non-metastasis) from bone metastasis was 0.995, 95% CI (0.986-1.000) ( Figure 1 (F) The above results indicate that secreted proteins TFF1 and TFF3 have good accuracy and specificity in predicting the early diagnosis of lung cancer patients.

[0070] Example 2 In vivo mouse experiment to detect the effect of TFF1 and TFF3 expression on lung cancer bone metastasis

[0071] 1. Establishment of lung cancer cell lines with stable high expression of TFF1 and TFF3 (PC9 TFF1 cells and PC9 TFF3 cells)

[0072] Generally, lentiviral vectors are used to prepare viral fluid (cell culture fluid containing viral plasmid particles, referred to as viral fluid). Transfection methods include liposome transfection and calcium phosphate transfection. The viral transfection experimental method is described using liposome transfection as an example.

[0073] Preparation before transfection: The number of cells is about 2.0-3.0×10 6 / 8mLCD human embryonic kidney cells (293FT cells) were plated in a 100×20mm cell culture dish; after 24 hours, the cell status was observed under a microscope to ensure that the cells were in good condition, and the transfection system was prepared: A solution and B solution, wherein A solution: psPAX2 (Addgene, Catalog: #12260) 6μg, pMD2.G (Addgene, Catalog: #12269) 4μg, TFF1 high expression plasmid (Weizhen Biotechnology Co., Ltd., Catalog: Catalog: #CH892913) or TFF3 high expression plasmid (Weizhen Biotechnology Co., Ltd., Catalog: #CH801849) 10μg, p3000 (Thermo, Catalog: #L3000-015) 15μL and Opti-MEM (Gibco, Catalog: #31985070) 500μL, B solution: 500μL Opti-MEM and 20μL lipofectamine 3000 (Thermo, Catalog: #L3000-015);

[0074] Transfection: Gently mix solution A and solution B, let stand at room temperature for 20 minutes, slowly and evenly add the mixed solution to the well-growing 293FT cells, and observe the cell status 4 to 6 hours after transfection;

[0075] Collect the virus solution: 24 hours after transfection, use a 10mL syringe to collect the virus solution, and filter it with a 0.45μm pore filter to remove cell debris. Collect the filtrate, which is the virus solution; at the same time, add fresh culture medium to the 293FT cells and continue to culture;

[0076] Preparation of infected cells: On the day of infection, the number of cells is about 1.5 × 10 5 After digestion and resuspension of the cells to be infected (PC9 cells) with 4mLCD, they were evenly spread to 25cm 2 In cell culture flasks;

[0077] Infect cells: Add 40 mg / mL of polybrene solution to the collected virus solution at a volume ratio of 1:1000, mix well, and infect PC9 cells after mixing. Collect the virus solution every 8 to 12 hours and infect for three consecutive days.

[0078] Screening positive cells: After three consecutive days of infection, Puromycin solution (10 mg / mL) was added to the culture medium at a volume ratio of 1:1000 to screen positive cells; the culture medium containing Puromycin was added to the infected cells to screen positive cells for 7 days, and the cell status and death were observed every day, and the medium was changed in time. When the positive cells grew stably, the positive cells were collected, and the cell RNA was extracted using the Trizol method, and the cell protein was extracted using SDS protein lysis buffer (Biyuntian, Catalog: #P0013G), and the efficiency of the stable overexpression cell line was verified by qRT-PCR and Western blot experiments; the cells after successful verification were expanded or frozen to obtain PC9 TFF1 cells and PC9 TFF3 cells;

[0079] 2. In vivo mouse experiments to detect the effects of TFF1 and TFF3 on lung cancer bone metastasis

[0080] Preparation of tumor cell suspension: digest and count PC9 TFF1 cells and PC9 TFF3 cells, use PC9 Vector cells as the control group, and resuspend the cells in PBS solution to a cell concentration of 5×10 5 / 100μL;

[0081] Ten 6- to 8-week-old balb / c-nu / nu mice were selected and randomly divided into three groups: a control group (injected with PC9 Vector cells), a PC9 TFF1 group (injected with PC9 TFF1 cells), and a PC9 TFF3 group (injected with PC9TFF3 cells). Each mouse was anesthetized by intraperitoneal injection of 100 μL 4% chloral hydrate. The tumor cell suspension was inoculated into the left ventricle of the mice, with 100 uL injected into each mouse. Be careful not to let air bubbles enter the syringe, and blood will spurt out of the syringe when the positioning is accurate. After injection, the mice were routinely cultured, and small animal in vivo imaging was used to continuously observe tumor metastasis for 2 to 6 weeks.

[0082] The results are as follows Figure 2 As shown, 4 weeks after tumor cell inoculation, the bone metastasis ability of lung cancer cell lines in PC9 TFF1 group and PC9TFF3 group was significantly enhanced compared with that in PC9 Vector group, that is, the bone metastasis ability of lung cancer cell lines with stable and high expression of TFF1 and TFF3 was significantly enhanced, indicating that TFF1 and TFF3 play an important role in the bone metastasis process of lung cancer.

[0083] The embodiments of the present invention are described in detail above in conjunction with the accompanying drawings, but the present invention is not limited to the above embodiments. Various changes can be made within the knowledge of ordinary technicians in the relevant technical field without departing from the purpose of the present invention. In addition, the embodiments of the present invention and the features in the embodiments can be combined with each other without conflict.

Claims

1. Use of a reagent for detecting biomarkers in the preparation of a product for diagnosis or early screening of lung cancer bone metastasis, wherein the biomarkers include TFF1 and / or TFF3.

2. The use according to claim 1, characterized in that: The reagents include reagents for quantitatively detecting biomarkers.

3. The use according to claim 2, characterized in that: The reagents for detecting biomarkers include reagents for detecting biomarkers at the gene level and / or protein level.

4. The use according to claim 3, characterized in that: The reagent for detecting biomarkers is a substance used in one or more detection techniques or methods selected from the following group: enzyme-linked immunosorbent assay, immunofluorescence, radioimmunoassay, immunocoprecipitation, immunoblotting, high performance liquid chromatography, capillary gel electrophoresis, near infrared spectroscopy, mass spectrometry, immunochemiluminescence, colloidal gold immunoassay, fluorescent immunochromatography, surface plasmon resonance, immuno-PCR or biotin-avidin technology.

5. The use according to any one of claims 1 to 4, characterized in that: The product includes at least one of a reagent, a test kit, a test paper, and a chip.

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