CN114062665A - Tracer particle marked target molecule and preparation method and application thereof - Google Patents

Tracer particle marked target molecule and preparation method and application thereof Download PDF

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Publication number
CN114062665A
CN114062665A CN202010780643.8A CN202010780643A CN114062665A CN 114062665 A CN114062665 A CN 114062665A CN 202010780643 A CN202010780643 A CN 202010780643A CN 114062665 A CN114062665 A CN 114062665A
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antigen
target molecule
virus antigen
virus
hepatitis
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CN114062665B (en
Inventor
郑锦龙
李瑞净
魏钟杰
黄荣生
岑静
覃丽芳
詹佩文
丁正辉
张耀东
陈一尘
庞土玲
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Fapon Biotech Inc
Guangdong Fapon Biotech Co Ltd
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Fapon Biotech Inc
Guangdong Fapon Biotech Co Ltd
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Abstract

The invention relates to the field of biological detection, and particularly provides a tracer particle labeled target molecule and a preparation method and application thereof. According to the preparation method of the tracer particle labeled target molecule, the target molecule and casein are mixed and then are combined with the tracer particle for reaction, and the tracer particle labeled target molecule is obtained. The method can improve the labeling state, enhance the binding capacity and stability of the target molecule and the tracer particle, and improve the activity and specificity in immune reaction.

Description

Tracer particle marked target molecule and preparation method and application thereof
Technical Field
The invention relates to the field of biological detection, in particular to a tracer particle labeled target molecule and a preparation method and application thereof.
Background
The immunochromatography technology is a rapid detection technology for qualitatively, semi-quantitatively or quantitatively detecting the content of an antigen or an antibody in a sample by utilizing an adsorption carrier and developing through tracer particles according to an immunological principle. Taking the detection of the antibody in the sample as an example, the antigen is firstly fixed in a certain zone of an adsorption carrier such as a nitrocellulose membrane, after one end of the dried nitrocellulose membrane is immersed in the sample (such as urine, saliva, serum), the sample will move forward along the membrane due to capillary action, when the dried nitrocellulose membrane moves to an area where the antigen is fixed, the corresponding antibody in the sample is specifically combined with the antigen, wherein the antigen is labeled by a tracer particle, and when the antigen-antibody combination reaction occurs, a certain color is displayed, so that the specific immunity detection is realized. Commonly used tracer particles include colloidal gold, latex, fluorescent microspheres, and the like.
In the preparation process of the immunochromatographic diagnostic reagent, the tracer particles are required to be combined with target molecules, namely, the tracer particle marking of the target molecules is realized. In the practical application process, for example, latex, colloidal gold, fluorescent microspheres, etc., all may have the problems that the binding force between the tracer particles and the target molecules is relatively weak, and the target molecules are easy to break. In the course of immunoreaction, if the target molecule in the sample has high affinity with the specific receptor/ligand, the target molecule and the trace particle are often broken, which leads to the problems of decreased immunoreaction activity, poor specificity, etc., for example, dead gold in the case of colloidal gold immunochromatography.
In view of the above, the present invention is particularly proposed.
Disclosure of Invention
The first purpose of the invention is to provide a preparation method of a tracer particle labeled target molecule.
A second object of the present invention is to provide a tracer particle labeled target molecule.
A third object of the invention is to provide the use of a tracer particle for labelling a target molecule.
The fourth purpose of the invention is to provide an immunochromatographic test strip.
In order to achieve the above purpose of the present invention, the following technical solutions are adopted:
a preparation method of a tracer particle labeled target molecule comprises the steps of mixing a target molecule with casein, and then carrying out a combined reaction with a tracer particle to obtain a tracer particle labeled target molecule;
the tracer particles comprise colloidal gold, colloidal selenium, colloidal silver, latex or fluorescent microspheres.
Further, the mass ratio of the target molecule to the casein is 1: (0.5-5);
optionally, the mass ratio of the target molecule to the casein is 1: (0.8-4);
optionally, the mass ratio of the target molecule to the casein is 1: (1-3.5);
optionally, the mass ratio of the target molecule to the casein is 1.5: (1.5-5).
Further, the target molecule is an antigen or an antibody;
further, the antigen or antibody is selected from the group consisting of antigens or antibodies related to the following immunoassay items: infectious disease immunodetection, myocardial marker immunodetection or tumor-associated marker immunodetection.
Further, the infectious diseases include viral infectious diseases, bacterial infectious diseases, fungal infectious diseases, chlamydia infectious diseases, mycoplasma infectious diseases, or parasitic infectious diseases;
optionally, the antigens associated with infectious diseases include HIV antigen, hepatitis A virus antigen, hepatitis B virus antigen, hepatitis C virus antigen, hepatitis D virus antigen, hepatitis E virus antigen, hepatitis G virus antigen, rubella virus antigen, human cytomegalovirus antigen, herpes simplex virus type 1 antigen, herpes simplex virus type 2 antigen, rabies virus antigen, human T lymphocyte leukemia virus antigen, dengue fever virus antigen, human papilloma virus antigen, West Nile virus antigen, forest encephalitis virus antigen, measles virus antigen, influenza virus antigen, parainfluenza virus antigen, varicella virus antigen, echovirus antigen, coxsackie virus antigen, encephalitis B virus antigen, coxsackie virus antigen, EB virus antigen, mumps virus antigen, treponema antigen, Borrelia burgdorferi antigen, Coxiella virus antigen, Epstein-Barr virus antigen, hepatitis B virus antigen, hepatitis C virus antigen, hepatitis D antigen, herpes simplex virus antigen, human T virus antigen, dengue virus antigen, chlamydia trachomatis antigen, chlamydia pneumoniae antigen, chlamydia psittaci antigen, ureaplasma urealyticum antigen, mycoplasma pneumoniae antigen, mycobacterium tuberculosis antigen, helicobacter pylori antigen, gonococcus antigen, plasmodium antigen, trypanosoma cruzi antigen or toxoplasma antigen;
optionally, the cardiac marker comprises troponin, myoglobin, creatine kinase isozyme, N-terminal brain natriuretic peptide precursor, cardiac fatty acid binding protein, D-dimer, lipoprotein-associated phospholipase a2, myeloperoxidase, or growth-stimulating expression gene 2 protein;
optionally, the tumor-associated markers comprise: gastrin releasing peptide precursor, carbohydrate antigen, cytokeratin 19, epididymin 4, alpha-fetoprotein, carcinoembryonic antigen, prostate specific antigen, squamous cell carcinoma antigen, or human prostate specific gene 1.
Further, the method also comprises a step of sealing after the target molecules are combined with the tracer particles, and then the tracer particle labeled target molecules are obtained.
The tracer particles prepared by the preparation method mark target molecules.
The tracer particle labeled target molecule is applied to immunodetection or preparation of an immunodetection reagent.
An immunochromatographic test strip comprises the tracer particle labeled target molecules.
Further, the immunochromatographic detection test paper comprises latex immunochromatographic detection test paper, colloidal gold immunochromatographic detection test paper, colloidal selenium immunochromatographic detection test paper, colloidal silver immunochromatographic detection test paper or fluorescent immunochromatographic detection test paper.
Compared with the prior art, the invention has the beneficial effects that:
according to the preparation method of the tracer particle labeled target molecule, the target molecule and casein are mixed and then are combined with the tracer particle for reaction, and the tracer particle labeled target molecule is obtained. In conventional cognition, the addition of unrelated proteins before the labeling of the tracer particle with the target molecule often competes for binding sites, resulting in reduced sensitivity. However, the inventors have found that the binding reaction between the target molecule and the tracer particle is carried out in an environment where casein exists, and the binding ability and stability of the target molecule and the tracer particle can be enhanced, so that the activity and specificity in the immune reaction can be improved, for example, the dead gold phenomenon disappears in the colloidal gold chromatography reaction, and an unexpected technical effect can be obtained. Particularly, in the prior art, a scheme that target molecules are difficult to combine with tracer particles and need indirect labeling exists, and the method for combining and reacting the target molecules and the casein and the tracer particles can achieve better effect than indirect labeling.
Detailed Description
Embodiments of the present invention will be described in detail below with reference to examples, but it will be understood by those skilled in the art that the following examples are only illustrative of the present invention and should not be construed as limiting the scope of the present invention. The examples, in which specific conditions are not specified, were conducted under conventional conditions or conditions recommended by the manufacturer.
Unless otherwise defined, technical and scientific terms used herein have the same meaning as is familiar to those skilled in the art. In addition, any methods or materials similar or equivalent to those described herein can also be used in the present invention.
The invention provides a preparation method of a tracer particle labeled target molecule, which comprises the steps of mixing a target molecule with casein, and then carrying out a combined reaction with the tracer particle to obtain the tracer particle labeled target molecule, wherein the tracer particle comprises colloidal gold, colloidal selenium, colloidal silver, latex or fluorescent microspheres.
In conventional cognition, the addition of unrelated proteins before the labeling of the tracer particle with the target molecule often competes for binding sites, resulting in reduced sensitivity. However, the inventors have found that an unexpected technical effect is achieved by performing a binding reaction between a target molecule and a tracer particle in an environment where casein exists, thereby improving the labeling state, enhancing the binding ability and stability of the target molecule and the tracer particle, and improving the activity and specificity in an immunoreaction, for example, the disappearance of a dead gold phenomenon in a colloidal gold chromatography reaction. Particularly, in the prior art, a scheme that target molecules are difficult to combine with tracer particles and need indirect labeling exists, and the method for combining and reacting the target molecules and the casein and the tracer particles can achieve better effect than indirect labeling.
It should be noted that the binding reaction between the target molecule and the tracer particle in the present invention may be a preparation process that is conventional in the art, and is not particularly limited herein, but the difference may be that casein is mixed into the target molecule, and then a mixture of the target molecule and the casein is bound to the tracer particle.
Colloidal gold is gold particles which are polymerized by chloroauric acid under the action of a reducing agent such as sodium citrate, white phosphorus, ascorbic acid, tannic acid and the like and have a certain size, and the gold particles become a stable colloidal state due to electrostatic action. Colloidal gold labeling is a process in which a polymer such as protein is adsorbed onto the surface of colloidal gold particles. The adsorption mechanism may be the negative charge on the surface of the gold colloid particles, which is combined with the positive charge group of the protein due to electrostatic adsorption. In addition, the colloidal selenium and the colloidal silver are also commonly used tracer particles, and the principle of labeling proteins is also the adsorption of colloidal metals.
The latex is a particle, and can be labeled by combining special groups on the surface of the latex, such as carboxyl groups, with amino groups of protein substances under the action of a catalyst, such as EDC.
The fluorescent microsphere is a tracing particle which is formed by adsorbing or embedding fluorescent dye into the particle through a physical adsorption method, a self-assembly method, a chemical bonding method, a copolymerization method, an embedding method and the like, has a diameter ranging from nanometer to micron, and can emit fluorescence under the excitation of an excitation light source.
It should be noted that labeling the target molecule with the tracer particle in the present invention corresponds to labeling the end in rapid diagnosis.
In a preferred embodiment, the mass ratio of target molecule to casein is: 1: (0.5-5), preferably 1: (0.8-4), more preferably 1: (1-3.5), more preferably 1.5: (1.5-5), specifically, the mass ratio may be, but is not limited to, 1: 0.5, 1:1, 1:1.5, 1:2, 1:2.5, 1:3, 1:3.5, 1:4, 1:4.5, or 1: 5. The ratio of the target molecule to the tracer particle can be used in the conventional amount in the art, and is not particularly limited herein.
In the present invention, the target molecule may be a substance labeled with a labeled particle, which is conventionally used in the art, and may be an antigen or an antibody. The antigen or antibody may be a relevant antigen or antibody for an immunoassay item commonly used in the art.
For example, the immunoassay items may be infectious disease immunoassays, myocardial marker immunoassays, or tumor-associated marker immunoassays.
The infectious disease may be a viral infectious disease, a bacterial infectious disease, a fungal infectious disease, a chlamydia infectious disease, a mycoplasma infectious disease, or a parasitic infectious disease.
The infectious disease associated antigen can be HIV antigen, hepatitis A virus antigen, hepatitis B virus antigen, hepatitis C virus antigen, hepatitis D virus antigen, hepatitis E virus antigen, hepatitis G virus antigen, rubella virus antigen, human cytomegalovirus antigen, herpes simplex virus 1 antigen, herpes simplex virus 2 antigen, rabies virus antigen, human T lymphocyte leukemia virus antigen, dengue fever virus antigen, human papilloma virus antigen, West Nile virus antigen, forest encephalitis virus antigen, measles virus antigen, influenza virus antigen, parainfluenza virus antigen, varicella virus antigen, echovirus antigen, coxsackievirus antigen, encephalitis B virus antigen, coxsackievirus antigen, EB virus antigen, mumps virus antigen, treponema antigen, Borrelia burgdorferi antigen, Cocci virus antigen, Epstein-Barr virus antigen, hepatitis B virus antigen, hepatitis C virus antigen, hepatitis D virus antigen, herpes simplex virus antigen, herpes virus antigen, human papillomavirus antigen, human infectious disease antigen, human papillomavirus antigen, human infectious disease antigen, Chlamydia trachomatis antigen, Chlamydia pneumoniae antigen, Chlamydia psittaci antigen, ureaplasma urealyticum antigen, Mycoplasma pneumoniae antigen, Mycobacterium tuberculosis antigen, helicobacter pylori antigen, gonococcus antigen, Plasmodium falciparum antigen, Trypanosoma cruzi antigen or Toxoplasma gondii antigen.
The cardiac marker may be troponin, myoglobin, creatine kinase isozyme, N-terminal brain natriuretic peptide precursor, cardiac fatty acid binding protein, D-dimer, lipoprotein-associated phospholipase a2, myeloperoxidase, or growth stimulating expression gene 2 protein.
The tumor associated marker may be a gastrin releasing peptide precursor, a carbohydrate antigen, cytokeratin 19, epididymin 4, alpha-fetoprotein, carcinoembryonic antigen, prostate specific antigen, squamous cell carcinoma antigen or human prostate specific gene 1.
In a preferred embodiment, the method further comprises a step of blocking the target molecule after the target molecule is bound to the tracer particle, and the tracer particle is obtained to label the target molecule. The blocking may be performed by a method conventional in the art, and is not particularly limited, for example, blocking with an inert protein such as bovine serum albumin.
The invention also protects the tracer particle marked target molecule prepared by the preparation method, and the tracer particle marked target molecule has good stability and high specificity and sensitivity for subsequent detection. It should be noted that the tracer particle labeled target molecule provided by the invention is used as a labeling end in specific applications such as rapid diagnostic technology. According to the difference between the tracer particles and the target molecules, the method can be widely applied to various fields, for example, when the target molecules are marked by colloidal gold, the method can be used for the specific detection of various substances such as electron microscope, optical microscope, flow cytometry, agglutination detection, immunoblotting, immunochromatography and the like.
The invention also protects the application of the tracer particle labeled target molecule in immunodetection or preparation of an immunodetection reagent.
The invention finally protects an immunochromatographic test paper which comprises the tracer particle labeled target molecule. Specifically, the test paper may be latex immunochromatographic test paper, colloidal gold immunochromatographic test paper, fluorescent immunochromatographic test paper, or the like.
The invention is further illustrated by the following specific examples, which, however, are to be construed as merely illustrative, and not limitative of the remainder of the disclosure in any way whatsoever.
Example 1 Indirect labeling procedure
1. Preparing 40nm colloidal gold with the concentration of 4 OD;
2. sucking 1ml of colloidal gold into a small glass bottle with the specification of 4 ml;
3. 0.2M K was added2CO3Shaking for 1min to adjust the pH of the colloidal gold;
4. adding 10 μ g GST monoclonal antibody and shaking for 1 min;
5. adding 10 μ l 10% BSA, sealing and shaking for 1 min;
6. transferring the uniformly mixed immune gold solution into a 1.5ml centrifuge tube, and placing the centrifuge tube in an ultracentrifuge at 1000rpm for 7 min;
7. carefully sucking the supernatant solution by using a pipette gun, and keeping a precipitate;
8. redissolving the precipitate to obtain 100 mul of concentrated gold;
9. adding 0.5 μ g of coupling protein (TP protein coupled GST) into the concentrated gold, and incubating for 15min to obtain the TP antigen (treponema pallidum antigen) of the colloidal gold.
Example 2 direct labeling procedure
1. Preparing 40nm colloidal gold with the concentration of 4 OD;
2. sucking 1ml of colloidal gold into a small glass bottle with the specification of 4 ml;
3. 0.2M K was added2CO3Shaking for 1min to adjust the pH of the colloidal gold;
4. adding 10 μ g of target protein (TP protein) and shaking for 1 min;
5. adding 10 μ l 10% BSA, sealing and shaking for 1 min;
6. transferring the uniformly mixed immune gold solution into a 1.5ml centrifuge tube, and placing the centrifuge tube in an ultracentrifuge at 9000rpm for 8 min;
7. carefully sucking the supernatant solution by using a pipette gun, and keeping a precipitate;
8. redissolving the precipitate to obtain 100. mu.L of concentrated gold, and obtaining the colloidal gold labeled TP antigen.
Example 3 direct labeling Casein Process
1. Preparing 40nm colloidal gold with concentration of 4 OD;
2. sucking 1ml of colloidal gold into a small glass bottle with the specification of 4 ml;
3. 0.2M K was added2CO3Shaking for 1min to adjust the pH of the colloidal gold;
4. mixing a target protein (TP protein) and casein (C) according to the mass ratio of 1.5:1.5 (scheme A), 1.5:3.5 (scheme B) and 1.5:5 (scheme C) respectively;
5. adding 10 μ g of the mixed target protein (TP + C) and shaking for 1 min;
6. adding 10 μ l 10% BSA, sealing and shaking for 1 min;
7. transferring the uniformly mixed immune gold solution into a 1.5ml centrifuge tube, and placing the centrifuge tube in an ultracentrifuge at 9000rpm for 8 min;
8. carefully sucking the supernatant solution by using a pipette gun, and keeping a precipitate;
9. redissolving the precipitate to obtain 100. mu.L of concentrated gold, and obtaining the colloidal gold labeled TP antigen.
Test example 1
Test strips were prepared using the colloidal gold-labeled TP antigens of examples 1 to 3, and the colloidal gold-labeled TP antigens obtained in examples 1 to 3 were diluted at a concentration of 15%; sucking 340 mul, and uniformly spreading on a 0.8cm × 30cm glass fiber membrane; freeze-drying for 2h using a freeze-dryer to obtain a gold pad for assembly; assembling the gold pad with other components (PVC base plate, NC film, absorbent paper, sample pad), and cutting into test strips of 2.5mm × 6cm with a slitter; adding 35 mul of sample on the test strip and interpreting; and recording the interpretation result.
The following samples were tested:
1. the test samples are 200 negative samples, and the activity and the positive detection number are counted, wherein the activity represents the attenuation from C1 to C9, and the results are shown in the following table:
Figure BDA0002618480310000091
2. detecting a sample as a positive quality control product: TP 2/TP 20/TP 60 (i.e. 2-fold, 20-fold, 60-fold dilution of the quality control material, respectively). The results are shown in the following table:
Figure BDA0002618480310000092
at present, the mainstream labeling mode of the TP antigen project is an indirect labeling method, namely, the labeling state of protein is more stable by coupling target protein to monoclonal antibody labeling, compared with the prior art, the protein is directly used for labeling the poor labeling state, the phenomena of sedimentation and dead gold occur, the color is more purple, the activity and the specificity are poorer than those of indirect labeling, but the casein and the target protein are mixed according to a certain proportion and then labeled, the labeling state can be improved, the original turbid and more purple state is changed into clear red, the specificity and the activity are improved and improved compared with those of indirect labeling, the color number of strong positive TP 2 of a quality control product is higher than that of indirect labeling 1C, the color number of medium positive TP 20 is higher than that of indirect labeling 1C, and the color number of weak positive TP 60 is equivalent to that of indirect labeling.
The random clinical test results showed that the test specificity of the random clinical sera of the direct marker + casein group was comparable to that of the indirect marker group.
Test example 2
1. Test strips were prepared according to the preparation procedures of examples 1-3 using HIV antigen (AIDS antigen) as the target protein to detect positive quality control HIV 20 (20-fold dilution of quality control), and the results are shown in the following table:
HIV-colloidal gold project Marking status Quality control HIV 20 visualization
Indirect labelling Clear red immortal gold C4
Direct marking Purple centrifuge dead gold solution C5
Direct labeling + Casein (1.5:3.5) Clear red immortal gold C4
2. Test strips were prepared according to the preparation procedures of examples 1 to 3 using HCV antigen as a target protein to detect positive quality control HCV 20 (20-fold dilution of quality control), and the results are shown in the following table:
HCV-colloidal gold project Marking status Quality control HCV 20 visualization
Indirect labelling Clear red immortal gold C5
Direct marking Purple centrifuge dead gold solution C6
Direct labeling + Casein (1.5:3.5) Clear red immortal gold C4
Example 4
The latex particles are used as marking particles, and the marking method is as follows:
1. preparing 100 mu L of latex particles, adding 900 mu L of 0.1M borate buffer solution with pH8.5, washing and resuspending;
2. adding 0.15mg EDC to activate the latex;
3. mixing target protein (HCV protein, namely hepatitis C antigen) and casein according to the mass ratio of 1.5: 3.5;
4. adding 10 μ g of the mixed target protein (HCV + C), and stirring overnight;
5. adding ethanolamine, sealing for 30min, centrifuging, and then re-suspending and concentrating to 100 mu L to obtain latex-labeled HCV antigen;
the test strips prepared by the latex-labeled HCV antigen detect positive quality control HCV 20 (the quality control is diluted by 20 times) and the positive detection number of 50 negative samples, the activity shows that the activity is enhanced from L1 to L10, and the results are shown in the following table:
HCV-latex program Specificity of Quality control HCV 20 visualization
Indirect labelling 1/50 L7
Direct marking 2/50 L6
Direct labeling + Casein (1.5:3.5) 0/50 L8
Example 5
The fluorescent microspheres are used as marking particles, and the marking method is as follows:
1. preparing 50 mu L of fluorescent microspheres (carboxyl polystyrene microspheres containing high-brightness rare earth dye), adding into 1mL of 0.01M MES buffer solution, and washing and resuspending;
2. adding 250 mu L of EDC, activating for 2h in a dark place, and washing and resuspending;
3. mixing a target protein (TP protein) and casein according to a mass ratio of 1.5: 3.5;
4. adding 10 mu g of the mixed target protein (TP + C), and reacting for 2 hours in a dark place;
5. respectively adding glycine and BSA (bovine serum albumin) and sealing for 30min in the dark, centrifuging and then carrying out heavy suspension concentration to 100 mu L to obtain a TP antigen marked by the fluorescent microspheres;
the test strip prepared by the fluorescent microsphere labeled TP antigen detects positive quality control TP 20 (the quality control product is diluted by 20 times) and the positive detection number of 50 negative samples, and the results are as follows:
TP-fluorescent microsphere project Specificity of Quality control TP 20 color development
Indirect labelling 1/50 8813
Direct marking 1/50 7744
Direct labeling + Casein (1.5:3.5) 0/50 9892
While particular embodiments of the present invention have been illustrated and described, it would be obvious that various other changes and modifications can be made without departing from the spirit and scope of the invention. It is therefore intended to cover in the appended claims all such changes and modifications that are within the scope of this invention.

Claims (10)

1. A preparation method of a tracer particle labeled target molecule is characterized in that the target molecule is mixed with casein and then is combined with a tracer particle for reaction to obtain the tracer particle labeled target molecule;
the tracer particles comprise colloidal gold, colloidal selenium, colloidal silver, latex or fluorescent microspheres.
2. The method according to claim 1, wherein the mass ratio of the target molecule to the casein is 1: (0.5-5);
optionally, the mass ratio of the target molecule to the casein is 1: (0.8-4);
optionally, the mass ratio of the target molecule to the casein is 1: (1-3.5);
optionally, the mass ratio of the target molecule to the casein is 1.5: (1.5-5).
3. The method of claim 1, wherein the target molecule is an antigen or an antibody.
4. The method according to claim 3, wherein the antigen or antibody is selected from the group consisting of antigens or antibodies related to the following immunoassay: infectious disease immunodetection, myocardial marker immunodetection or tumor-associated marker immunodetection.
5. The method according to claim 4, wherein the infectious disease includes a viral infectious disease, a bacterial infectious disease, a fungal infectious disease, a chlamydia infectious disease, a mycoplasma infectious disease, or a parasitic infectious disease;
optionally, the antigens associated with infectious diseases include HIV antigen, hepatitis A virus antigen, hepatitis B virus antigen, hepatitis C virus antigen, hepatitis D virus antigen, hepatitis E virus antigen, hepatitis G virus antigen, rubella virus antigen, human cytomegalovirus antigen, herpes simplex virus type 1 antigen, herpes simplex virus type 2 antigen, rabies virus antigen, human T lymphocyte leukemia virus antigen, dengue fever virus antigen, human papilloma virus antigen, West Nile virus antigen, forest encephalitis virus antigen, measles virus antigen, influenza virus antigen, parainfluenza virus antigen, varicella virus antigen, echovirus antigen, coxsackie virus antigen, encephalitis B virus antigen, coxsackie virus antigen, EB virus antigen, mumps virus antigen, treponema antigen, Borrelia burgdorferi antigen, Coxiella virus antigen, Epstein-Barr virus antigen, hepatitis B virus antigen, hepatitis C virus antigen, hepatitis D antigen, herpes simplex virus antigen, human T virus antigen, dengue virus antigen, chlamydia trachomatis antigen, chlamydia pneumoniae antigen, chlamydia psittaci antigen, ureaplasma urealyticum antigen, mycoplasma pneumoniae antigen, mycobacterium tuberculosis antigen, helicobacter pylori antigen, gonococcus antigen, plasmodium antigen, trypanosoma cruzi antigen or toxoplasma antigen;
optionally, the cardiac marker comprises troponin, myoglobin, creatine kinase isozyme, N-terminal brain natriuretic peptide precursor, cardiac fatty acid binding protein, D-dimer, lipoprotein-associated phospholipase a2, myeloperoxidase, or growth-stimulating expression gene 2 protein;
optionally, the tumor-associated markers comprise: gastrin releasing peptide precursor, carbohydrate antigen, cytokeratin 19, epididymin 4, alpha-fetoprotein, carcinoembryonic antigen, prostate specific antigen, squamous cell carcinoma antigen, or human prostate specific gene 1.
6. The method of any one of claims 1-5, wherein the step of blocking the target molecule after binding the target molecule to the tracer particle comprises obtaining the tracer particle labeled target molecule.
7. The labeled target molecule prepared by the method according to any one of claims 1 to 6.
8. Use of a tracer particle labeled target molecule according to claim 7 in an immunoassay or in the preparation of an immunoassay reagent.
9. An immunochromatographic test strip comprising the labeled target molecule of claim 7 labeled with the tracer particle.
10. The immunochromatographic test strip of claim 9, wherein the immunochromatographic test strip comprises a latex immunochromatographic test strip, a colloidal gold immunochromatographic test strip, a colloidal selenium immunochromatographic test strip, a colloidal silver immunochromatographic test strip or a fluorescent immunochromatographic test strip.
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