CN106632617B - Antigen for detecting anti-human papilloma virus antibody and related immunoassay kit - Google Patents
Antigen for detecting anti-human papilloma virus antibody and related immunoassay kit Download PDFInfo
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Abstract
The invention relates to a Human Papilloma Virus (HPV) oncogene protein serving as an antigen for detecting an antibody resisting Human Papilloma Virus (HPV), wherein the sequence of the antibody is shown as SEQ ID NO. 4. Also provides an immunoassay kit comprising the antigen. And the application of the antigen and the kit. The antigen for detecting the anti-HPV antibody can detect the HPV antibody in blood or saliva with high sensitivity, high specificity and high stability, thereby bringing basis for doctors to accurately diagnose cervical intraepithelial neoplasia and cervical cancer transformed by HPV oncogene of patients, and then taking treatment in time to prevent cancer occurrence or cancer diffusion and relieve pain of the patients in time.
Description
The application is a divisional application of an invention application with application date of 2013, 2 and 22 months, application number of 201310056516.3, and invention name of an antigen for detecting anti-human papilloma virus antibody and a related immunoassay kit.
Technical Field
The invention relates to the technical field of antibody detection, in particular to the technical field of anti-Human Papilloma Virus (HPV) antibody detection, specifically to an antigen for detecting anti-human papilloma virus antibody and a related immunodetection kit, and especially can be used for high-risk Human Papilloma Virus (HPV) subtypes HPV16 and HPV18 which are popular in China.
Background
Cervical cancer is the second most common female malignancy, for which Human Papillomavirus (HPV) is a prerequisite. Although HPV infections are common in humans, such as: the infection rate of women in the high cervical cancer incidence area is still up to 20.8 percent after the age of 35 years. However, over 80% of women who are HPV DNA positive do not progress to cancer or precancerous lesions. Thus, HPV DNA detection is deficient in being unable to distinguish between a transient infection and an infection that may cause cervical lesions, resulting in a higher false positive rate. A large number of researches show that the expression product of the HPV oncogene (oncoprotein) has high oncogenic activity, so that the biomarkers can be used as more accurate and specific cervical cancer prediction indexes than HPV DNA. The expression level of high-risk HPV E2, E6 and E7 oncoprotein (oncoprotein) is directly related to different stages of cervical cancer precancerous lesion. By utilizing the characteristic, a brand-new cervical cancer molecular screening index can be constructed.
Research on HPV carcinogenic mechanism shows that early expression proteins E6 and E7 of high-risk HPV play an important role in the generation of cervical cancer. HPV E6 and E7 proteins are generally over-expressed in cervical epithelial cells of patients with high-grade cervical lesions and cancers, and E6 and E7 oncoproteins which are continuously expressed and over-expressed can interact with host cell regulatory proteins (such as P53 and Rb proteins and the like) to inactivate tumor suppressors, so that cells are immortalized and malignantly transformed.
In addition to HPV DNA detection and detection of HPV oncoproteins, HPV antibodies in blood (body fluids) also demonstrate a great correlation with HPV infection and history of cervical cancer development. HPV L1 blood antibodies can be detected using standard ELISA methods, providing information on HPV infection. anti-HPV antibodies are induced by HPV as the time for HPV infection is prolonged, so that anti-HPV antibodies in serum can be detected. Wikstrom et al detected serum IgG antibodies to HPV6 and HPV11 type capsid proteins in 47 men with current condyloma acuminatum and 32 men with a history of condyloma acuminatum and compared them with 205 men with no history of genital condyloma acuminatum, showed that IgG antibodies against HPV6 type in serum were 35% in men with a history of condyloma acuminatum, 27% in current condyloma acuminatum and 10% in the control group. This indicates that anti-HPV 6IgG antibodies were present at a later time in condyloma acuminatum, while IgG positivity also reflects previous infection with the virus.
Stone et al reported the Infection rate study of 7218 cases of HPV-16 in the general population of the United states using this method, and the results showed a female positive rate of 17.9% and a male positive rate of 7.9% (Stone, K.M., et al, prediction of human Papilomavir Type 16Infection in the United states 2002.the journal Infection Diseases 2002; 186: 1396-. The HPV L1 antibody ELISA method has been widely adopted in the development and clinical trials of HPV vaccine (Merck Gardasil preventive vaccine). Luevano et al performed High Throughput assay with protein chips on serum samples from cervical cancer patients (Luevano, M., et al, High-Throughput Profiling of the Human Immune response Against third Human Immune proteins, micro microorganisms, 2010. virology.2010; 405(1): 31-40), analyzing antibody Profiling of a total of 54604 proteins of the 13 HPV Types, and showed strong correlation of antibodies to oncogenic proteins in blood with cervical cancer, High-grade lesions, low-grade lesions, and asymptomatic normal persons (Lueano, M., et al, High-Throughput Profiling of the Human Immune Responses Against third Human Immune proteins, micro microorganisms, 2010. 31-40).
Therefore, in order to achieve high sensitivity, high specificity and high stability of HPV antibody detection, it is necessary to provide a new antigen for detecting anti-human papillomavirus antibodies.
Disclosure of Invention
In order to solve the above problems, the present invention provides an antigen for detecting anti-HPV antibody and a related immunoassay kit, wherein the antigen for detecting anti-HPV antibody can detect HPV antibodies in blood or saliva with high sensitivity, high specificity and high stability, so as to provide a basis for doctors to accurately diagnose cervical intraepithelial neoplasia and cervical cancer of patients, and further to take timely treatments to prevent cancer occurrence or cancer diffusion and relieve pain of patients.
In order to achieve the above object, in a first aspect of the present invention, there is provided an antigen for detecting an anti-human papillomavirus antibody, characterized in that the antigen for detecting an anti-human papillomavirus antibody is selected from the group consisting of an amino acid sequence shown in SEQ ID NO. 1, an amino acid sequence shown in SEQ ID NO.2, an amino acid sequence shown in SEQ ID NO.3, an amino acid sequence shown in SEQ ID NO.4, an amino acid sequence shown in SEQ ID NO. 5, an amino acid sequence having 80% homology with the amino acid sequence shown in SEQ ID NO. 1, an amino acid sequence having 80% homology with the amino acid sequence shown in SEQ ID NO.2, an amino acid sequence having 80% homology with the amino acid sequence shown in SEQ ID NO.3, an amino acid sequence having 80% homology with the amino acid sequence shown in SEQ ID NO.4, and an amino acid sequence shown in SEQ ID NO. 5 One or more of amino acid sequences with 80% homology.
Preferably, the m +2 th, m +3 th, m +5 th, m +6 th and m +8-m +12 th amino acids of the antigen for detecting anti-human papilloma virus antibody are Asp, Ser, Glu, Asp, Glu, Ile, Asp and Gly in sequence, wherein m is more than or equal to 0.
Preferably, the antigen for detecting anti-human papillomavirus antibody is obtained by chemical synthesis or biological engineering method. Example (c): recombinant proteins, fusion proteins.
In a second aspect of the present invention, there is provided an immunoassay kit characterized by comprising the above-mentioned antigen for detecting an anti-human papillomavirus antibody.
Preferably, the immunoassay kit further comprises a carrier on which the antigen for detecting an anti-human papillomavirus antibody is coated.
Preferably, the immunoassay kit further comprises a labeled secondary antibody against human papillomavirus antibody. The label may be horseradish peroxidase, alkaline phosphatase, fluorescent molecule FITC (or other fluorescent label), chemiluminescent detection. The detection method can be qualitative and quantitative analysis by a color development method, a fluorescence method, chemiluminescence and an electrochemiluminescence method.
More preferably, the labeled secondary antibody against human papillomavirus antibody is a secondary antibody against human papillomavirus antibody labeled with horseradish peroxidase.
In a third aspect of the present invention, there is provided the use of the above antigen for detecting anti-human papillomavirus antibodies or the above immunoassay kit for detecting anti-human papillomavirus antibodies in human body fluids.
Preferably, the body fluid is blood or saliva.
The invention has the beneficial effects that: the antigen for detecting the anti-human papilloma virus antibody is selected from one or more of an amino acid sequence shown by SEQ ID NO. 1, an amino acid sequence shown by SEQ ID NO.2, an amino acid sequence shown by SEQ ID NO.3, an amino acid sequence shown by SEQ ID NO.4, an amino acid sequence shown by SEQ ID NO. 5, an amino acid sequence with 80 percent homology with the amino acid sequence shown by SEQ ID NO. 1, an amino acid sequence with 80 percent homology with the amino acid sequence shown by SEQ ID NO.2, an amino acid sequence with 80 percent homology with the amino acid sequence shown by SEQ ID NO.3, an amino acid sequence with 80 percent homology with the amino acid sequence shown by SEQ ID NO.4 and an amino acid sequence with 80 percent homology with the amino acid sequence shown by SEQ ID NO. 5, after a patient is infected by virus, the specific antibodies IgM, IgA or IgG molecules which appear in serum and oral saliva can be detected in a high-specificity, high-sensitivity and high-specificity manner, so that the basis can be brought to doctors for accurately diagnosing cervical intraepithelial neoplasia and cervical cancer of the patient, and then the treatment can be timely adopted to prevent cancer from happening or spread of cancer and relieve the pain of the patient in time.
Detailed Description
In order that the technical contents of the present invention can be more clearly understood, the following specific examples are specifically enumerated. However, the specific embodiments are merely illustrative, and not restrictive of the invention.
Materials and methods
1. Reagent and medicine
Horse radish peroxidase labeled Goat Anti-Human (Goat Anti-Human IgG, FC) was purchased from Prime Product # 31413.
Sodium chloride, potassium chloride, disodium hydrogen phosphate, and potassium dihydrogen phosphate were purchased from national pharmaceutical group chemical agents, ltd. Tween-20 was purchased from AMRESCO. Fetal bovine serum was purchased from Beijing Yuan Hengjin Ma Biotechnology development, Inc.
2. Instrument and consumable
Electronic balances (mertler-toledo instruments shanghai ltd); 85-1 constant temperature magnetic stirrer (Jie Rui Er electric appliances Co., Ltd., Kingsu Jintan city); a MULTISKAN MK3 microplate reader (Thermo Co.); ultrapure water preparation apparatus (MILLPORE corporation, usa); GZX-9240MBE digital display blast drying oven (Shanghai Boxun industry Co., Ltd.); WellWASK4MK2 plate washer (available from Thermo corporation); microplate (COSTAR corporation); single-channel, multi-channel pipettors (eppendorf, Germany).
3. Primary solution system
Coating buffer (PBS phosphate solution): 0.2g of monopotassium phosphate, 2.9g of 12-water disodium hydrogen phosphate, 8.0g of sodium chloride and 0.2g of potassium chloride, and distilled water is added to 1L for storage at 4 ℃.
Wash buffer (PBS-Tween 20, 0.1%): adding Tween-201 mL into 1L of dilution buffer solution (PBS), uniformly stirring, and adjusting the pH value to 7.0-7.2.
Blocking solution: 5% milk-PBS-Tween
Serum dilutions: 10% FBS-PBS-Tween
TMB display: amresc, CAT No.64285730, mixed in equal volumes with pre-TMB base color development solution A and B.
Substrate color development solution a: 13.6g of sodium acetate, 1.6g of citric acid, 0.3ml of hydrogen peroxide (30 percent) and distilled water added to 500 ml.
o substrate color B solution: disodium ethylenediaminetetraacetate 0.2g citric acid 0.95g glycerol 50ml tetramethylbenzidine 0.2g distilled water to 500 ml.
Stop solution (2mol/L H2SO 4): 178.3mL of double distilled water, 21.7mL of concentrated sulfuric acid (98%) was added dropwise.
4. Establishment of Indirect ELISA method
1) The HPV polypeptide antigen HPV16-1/HPV16-2/HPV16-3/HPV18-1/HPV18-2 was diluted with PBS at a concentration of 4ug/ml, coated on costar plates at 100 ul/well and left overnight at 4 ℃.
2) PBS was washed once and patted dry.
3) 5% milk powder-PBS was added to block, 250 ul/well, 37 degrees for 3 hours.
4) PBST was washed once and patted dry.
5) 10% FBS-PBST dilution serum 1: diluting with 50 deg.C, and mixing with oscillator.
6) Diluted serum, blank, negative and positive controls, 50 ul/well, 37 degrees were added and incubated for 1 hour.
7) 0.1% tween-PBS was washed 5 times and patted dry.
8) Goat anti-human was labeled with horseradish peroxidase, diluted with 10% FBS-PBST (1: 40000) Incubate 100 ul/well at 37 ℃ for 30 min.
9) 0.1% tween-PBS was washed 5 times and patted dry.
10) TMB color development, wherein TMB solution A and B solution are mixed in equal volume, 100 ul/hole, and 37 ℃ for 20 min.
11)2M H2SO4The wells were stopped at 50 ul/well and read at 450nm using a microplate reader.
5. The kit comprises the following components:
the kit consists of the following parts:
1. costar enzyme label plate coated with antigen for detecting anti-human papilloma virus antibody
2. Horse radish peroxidase labeled goat anti-human tube (10ul)
3.10 PBST bottle (10ml)
One bottle (6ml) of TMB color development liquid A and one bottle (6ml) of B liquid
5. Stopping solution bottle (6ml)
6. Envelope antigens
The polypeptide antigen is as follows
| HPV16-1 | KCDSTLRLCVQSTHVDIRTLE | SEQ ID NO：1 |
| HPV16-2 | PTLHEYMLDLQPETTDLYCYEQLNDSSEEE | SEQ ID NO：2 |
| HPV16-3 | LNDSSEEEDEIDGPAGQAEPDRAH | SEQ ID NO：3 |
| HPV18-1 | IDGVNHQHLPARRAEPQR | SEQ ID NO：4 |
| HPV18-2 | SDSEEENDEIDGVNHQHLPARRAEPQRH | SEQ ID NO：5 |
Second, the detection result
The kit selects 5 polypeptides (SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO: 5) as coating selection, and detects serum by an ELISA method.
1. Detecting the serum of the healthy human by using an ELISA (enzyme-linked immunosorbent assay) method by using an antigen-coated and sealed ELISA plate, wherein the result of the detected original data is as follows;
table one: HPV blood ELISA detection normal human serum result table
2.2 polypeptides (SEQ ID NO:3, SEQ ID NO: 5) are selected as an enzyme label plate which is coated, selected and sealed, and normal human serum is detected by an ELISA method, wherein the result of the detected original data is as follows;
table two: HPV blood ELISA detection normal human serum result table
3. 5 polypeptides (SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO: 5) are selected as enzyme label plates coated, selectively coated and sealed to detect serum of a patient by an ELISA method, the pathological diagnosis result of the patient is shown in the third table, wherein part of patients are subjected to HC2 pathological examination, and the result is as follows:
table three: HPV blood ELISA test patient serum result table
4. 3 polypeptides (SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO: 5) are selected as enzyme label plates coated, selectively coated and sealed to detect the serum of a patient by an ELISA method, the pathological diagnosis result of the patient is shown in the fourth table, wherein part of the patients are subjected to HC2 pathological examination, and the results are as follows:
table four: HPV blood ELISA test patient serum result table
Third, data analysis
1. According to the detection value of normal person, the average value, standard deviation and CUTOFF value can be calculated. The reaction OD values of 44 normal human serums on HPVE7 polypeptide in the table I are respectively averaged, and the standard deviation SD is calculated, when the kit determines the CUTOFF value, the average value of the normal human sera plus twice the standard deviation is selected as the judgment standard of HPV negative and positive, and the result is as follows:
the Cutoff calculation formula is as follows: cutoff is the mean OD +2 SD standard deviation
1.1 according to the results in Table I, the statistical analysis of the results of the serum test of 5 polypeptides HPVE7(SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO: 5) in normal humans showed the following results:
table five: data table for 5 polypeptide serum ELISA detection of HPV E7
| SEQ ID NO：1 | SEQ ID NO：2 | SEQ ID NO：3 | SEQ ID NO：4 | SEQ ID NO：5 | |
| Mean value of | 0.292 | 0.279 | 0.382 | 0.301 | 0.383 |
| Standard deviation of the mean | 0.098 | 0.100 | 0.111 | 0.076 | 0.111 |
| cutoff | 0.487 | 0.480 | 0.604 | 0.452 | 0.605 |
1.2 according to the results of Table two, the results of statistical analysis of the results of the detection of normal human sera by HPVE7(SEQ ID NO:3 and SEQ ID NO: 5)2 polypeptides are shown in the following table:
table six: data table for ELISA detection of 2 polypeptide sera of HPV E7
1.3 according to the results of Table five and Table six, comparative analysis gave the amino acid sequence of SEQ ID NO:3 and SEQ ID NO:5 the 2 polypeptides have consistent cutoff values in different experiments.
2. And judging whether the detected patient serum HPV is negative or positive according to the cutoff value, wherein the patient serum HPV is considered to be positive if the cutoff value is exceeded, and the patient serum HPV is considered to be infected if only one polypeptide is detected to be positive, and meanwhile, comparing the result with the HC2 diagnosis result.
2.1 in the tested patient serum 291 cases have HC2 diagnosis results, and in the case 84 cases with positive pathological diagnosis, in the pathological positive patient group (clinical diagnosis "gold standard"), the serum antibody detection result is compared with the HC2 molecule detection result (see Table 7), and the following table is compared with the HPV detection result of the kit: (statistically positive in the present test result includes (i) a positive result in either of the two antigen test results and (ii) a positive result in both of the two antigen test results)
TABLE VII: from the results in Table three, the serum ELISA diagnosis for the patient is compared with the HC2 diagnosis
("+" indicates positive result, "-" indicates negative result)
Table eight: from the results in Table four, the serum ELISA diagnosis for the patients is compared with the HC2 diagnosis
("+" indicates positive result, "-" indicates negative result)
The results of HC2 and the results of the blood ELISA test of the seven results in the following Table were analyzed by people statistics:
table nine: HC2 result and blood ELISA detection accuracy rate number and proportion comparison table
Other gynecological diseases mainly include pelvic inflammatory disease, ovarian cyst, hysteromyoma, etc.
The results of HC2 and the results of the blood ELISA test of the results of Table eight were analyzed by people statistics, and the results are shown in the following table:
TABLE Ten: HC2 result and blood ELISA detection accuracy rate number and proportion comparison table
Other gynecological diseases mainly include pelvic inflammatory disease, ovarian cyst, hysteromyoma, etc.
2.2 there are 10 cases of serum in the patient's serum that has no HC2 detection report, but the pathology report is CIN and cervical carcinoma, the result of HPV detection, patient number and blood detection positive rate comparison result of this kit are shown in the table.
Table eleven: for patients without HC2 diagnosis but with pathological conclusion, the results of ELISA test with HPV blood are summarized
("+" indicates positive result, "-" indicates negative result)
Table twelve: CIN without HC2 result, number of people for ELISA detection of patients with cervical cancer and cervical lesion
| Pathology of disease | Number of people (10) | SEQ ID NO:1 positive | SEQ ID NO:3 positive | SEQ ID NO:5 positive |
| CINⅠ | 2 | 2 | 1 | 1 |
| CINⅡ，Ⅲ | 7 | 4 | 5 | 5 |
| Cervical cancer | 1 | 0 | 1 | 1 |
2.3 statistics of the number of patients tested by ELISA and the results of positive ratio of all patients with CINIII and cervical cancer are shown in Table thirteen.
Table thirteen: the number of patients with CIN, cervical cancer and cervical lesion in blood ELISA detection and the accuracy rate are proportional
The polypeptide is selected by the inventor and is detected and verified in a patient sample and a normal human sample, and the GST-HPV16-E7 and GST-HPV118E7 full-length fusion protein (sequences shown as SEQ ID NO: 6 and SEQ ID NO: 7 respectively) is directly adopted to detect serum, so that the effect is poor, and the following experimental data are shown. Or
Fourth, the experimental materials and ELISA method are the same as the first point, except that the polypeptide (SEQ ID NO:3, SEQ ID NO: 5) is used for detection, GST-HPV16-E7 and GST-HPV118E7 full-length fusion protein are also used as detection antigens, the concentration is 2ug/ml when the GST-HPV16-E7 and GST-HPV18-E7 full-length fusion proteins are diluted by PBS, and the detection results are as follows:
1. according to the above experimental procedures, some of the patient serum and normal human serum were measured, and the results and pathological reports of the patients are shown in the following table:
fifth, analysis of the experiment
1. The results of the assay using the full-length protein were compared to the results of the prior combined assay using the HPV polypeptides SEQ ID:3 and SEQ ID:5 as follows:
to summarize: to summarize: the full-length fusion proteins of HPV16-E7 and HPV18-E7 have certain positive patients with missed detection and poorer sensitivity than polypeptide detection when detecting CIN and cervical cancer patients. The kit has a certain false positive rate when detecting patients with common gynecological inflammation and common people, and the sensitivity is slightly worse than that of polypeptide detection. Therefore, the full-length proteins of HPV16-E7 and HPV18-E7 are not well-suited for detection of CIN, cervical cancer and the like.
The blood enzyme-linked immunosorbent assay (ELISA) provided by the invention is based on the immune biochemical principle, and provides an antigen-antibody detection method with high sensitivity, high specificity and high stability. After infection with a virus, the patient may present specific antibodies in serum and oral saliva, IgM, IgA or IgG molecules. The specific anti-HPV antibody molecules in the serum or saliva of a patient are detected by ELISA (enzyme-linked immunosorbent assay). The invention analyzes the HPV high-risk HPV16 and HPV18 gene sequences, uses bioinformatics analysis to theoretically predict antigenicity, and uses an experimental method to screen and prove the significance of clinical application. By preparing a plurality of high-purity antigen polypeptides and repeatedly screening and verifying the prepared antigens by using serum of healthy people and serum of infected people in China, the target antigens HPV16-1, HPV16-2, HPV16-3, HPV18-1 and HPV18-2 with high sensitivity, high specificity and high stability are obtained. And proves that the polypeptide antigens can maintain the immunoreaction activity by being combined with the surface of a certain solid phase carrier. In the measurement, the specimen to be tested is reacted with the antigen on the surface of the solid carrier. The antigen and antibody complex formed on the solid phase carrier is separated from other matters by washing, and finally the enzyme bound on the fixed carrier has a certain ratio to the detected matter in the specimen. After the substrate for the enzyme reaction is added, detection is carried out by color development, so that qualitative or quantitative analysis can be carried out according to the shade of the color reaction. Compared with the DNA detection (HC2, CareHPV), the blood ELISA detection kit has the advantages of uniform sampling, non-invasiveness, quicker and simpler operation; the cost is low, the popularization is easy, and the method is more suitable for the Chinese market; false positive and false negative brought by PCR method can be avoided, and accuracy and precision of detection result are greatly improved; compared with the traditional pap smear, acetic acid staining VIA and colposcopy, the method is more accurate, easier and faster. The ELISA detection method can be used as a tool for large-scale screening for early screening of cervical cancer through optimization of various parameters and experimental procedures.
In conclusion, the antigen for detecting the anti-human papilloma virus antibody can detect the HPV antibody in blood or saliva with high sensitivity, high specificity and high stability, thereby bringing the basis for doctors to accurately diagnose cervical intraepithelial neoplasia and cervical cancer of patients, and then taking treatment in time to prevent cancer occurrence or cancer diffusion and relieve the pain of the patients in time.
In this specification, the invention has been described with reference to specific embodiments thereof. It will, however, be evident that various modifications and changes may be made thereto without departing from the broader spirit and scope of the invention. The description is thus to be regarded as illustrative instead of limiting.
Claims (12)
1. An antigen combination for detecting anti-human papillomavirus antibodies, wherein said antigen combination consists of the following antigens: an antigen with an amino acid sequence shown as SEQ ID NO.4, an antigen with an amino acid sequence shown as SEQ ID NO.3 and an antigen with an amino acid sequence shown as SEQ ID NO. 2.
2. An antigen combination for detecting anti-human papillomavirus antibodies, wherein said antigen combination consists of the following antigens: an antigen with an amino acid sequence shown as SEQ ID NO. 1, an antigen with an amino acid sequence shown as SEQ ID NO.2, an antigen with an amino acid sequence shown as SEQ ID NO.3, an antigen with an amino acid sequence shown as SEQ ID NO.4 and an antigen with an amino acid sequence shown as SEQ ID NO. 5.
3. An antigen combination for detecting anti-human papillomavirus antibodies, wherein said antigen combination consists of the following antigens: an antigen with an amino acid sequence shown as SEQ ID NO.2, an antigen with an amino acid sequence shown as SEQ ID NO.3, an antigen with an amino acid sequence shown as SEQ ID NO.4 and an antigen with an amino acid sequence shown as SEQ ID NO. 5.
4. An immunoassay kit comprising the antigen combination for detecting anti-human papillomavirus antibodies according to any one of claims 1 to 3.
5. The immunoassay kit of claim 4, further comprising a carrier on which the antigen is coated.
6. The immunoassay kit of claim 4, further comprising a secondary antibody against an antibody against human papilloma virus labeled with a label.
7. The immunoassay kit of claim 6, wherein the label is selected from the group consisting of: horseradish peroxidase, alkaline phosphatase, fluorescent labels and chemiluminescent labels.
8. Use of the antigen combination for detecting anti-human papillomavirus antibodies according to any one of claims 1 to 3 in the preparation of a reagent for the detection of human papillomavirus antibodies.
9. The use of claim 8, wherein the agent is also for diagnosing cervical intraepithelial neoplasia or cervical cancer.
10. Use of the antigen combination for detecting anti-human papillomavirus antibodies according to any one of claims 1 to 3 or of the immunoassay kit according to claim 4, for the preparation of a kit for the detection of anti-human papillomavirus antibodies in human body fluids.
11. Use according to claim 10, wherein the body fluid is blood or saliva.
12. The use according to claim 10, wherein the kit for detecting anti-human papillomavirus antibodies in human fluids is also used for the diagnosis of cervical intraepithelial neoplasia or cervical cancer.
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0594613A1 (en) * | 1990-05-11 | 1994-05-04 | Ferring AB | Synthetic peptides of human papillomaviruses 1, 6, 11, 16, 18, 31, 33 and 56, useful in immunoassay for diagnostic purposes |
| CN1775802A (en) * | 2004-11-17 | 2006-05-24 | 宋硕 | Human papillomavirus immunogenic polypeptide, and its preparing method and use |
| CN103254307A (en) * | 2012-02-16 | 2013-08-21 | 艾托金生物医药（苏州）有限公司 | HPV18E7 monoclonal antibody, hybridoma cell strain and application |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| ATE234860T1 (en) * | 1991-07-13 | 2003-04-15 | Dade Behring Marburg Gmbh | USE OF HPV-16 E6 AND E7 GENE OBTAINED PEPTIDES FOR DIAGNOSTIC PURPOSE |
| US7026443B1 (en) * | 1999-12-10 | 2006-04-11 | Epimmune Inc. | Inducing cellular immune responses to human Papillomavirus using peptide and nucleic acid compositions |
| CN1433428A (en) * | 2000-04-05 | 2003-07-30 | 冲击诊断公司 | Immunological methodology for discerning human papillomavirus |
| US7314630B2 (en) * | 2005-01-07 | 2008-01-01 | Yao-Xiong Hu | Compounds and methods of early diagnosis of cervical cancer and genital condyloma with HPV, CHSP60 tumor suppressor H-Ras, K-Ras and PTEN derived peptides modified |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP0594613A1 (en) * | 1990-05-11 | 1994-05-04 | Ferring AB | Synthetic peptides of human papillomaviruses 1, 6, 11, 16, 18, 31, 33 and 56, useful in immunoassay for diagnostic purposes |
| CN1775802A (en) * | 2004-11-17 | 2006-05-24 | 宋硕 | Human papillomavirus immunogenic polypeptide, and its preparing method and use |
| CN103254307A (en) * | 2012-02-16 | 2013-08-21 | 艾托金生物医药（苏州）有限公司 | HPV18E7 monoclonal antibody, hybridoma cell strain and application |
Non-Patent Citations (1)
| Title |
|---|
| Human Papillomavirus Type 18 E6 and E7 Antibodies in Human Sera: Increased Anti-E7 Prevalence in Cervical Cancer Patients;CONRAD BLEUL 等;《JOURNAL OF CLINICAL MICROBIOLOGY》;19910831;第29卷(第8期);第1581-1582页，附图1-2 * |
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