CN1152053C - Method for induction and prepn. of epitope idiosyncratic monoclonal antibody by using epitope polypeptide - Google Patents
Method for induction and prepn. of epitope idiosyncratic monoclonal antibody by using epitope polypeptide Download PDFInfo
- Publication number
- CN1152053C CN1152053C CNB991238087A CN99123808A CN1152053C CN 1152053 C CN1152053 C CN 1152053C CN B991238087 A CNB991238087 A CN B991238087A CN 99123808 A CN99123808 A CN 99123808A CN 1152053 C CN1152053 C CN 1152053C
- Authority
- CN
- China
- Prior art keywords
- epitope
- polypeptide
- monoclonal antibody
- polypeptides
- epitope polypeptide
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Fee Related
Links
- 229920001184 polypeptide Polymers 0.000 title claims abstract description 25
- 102000004196 processed proteins & peptides Human genes 0.000 title claims abstract description 25
- 108090000765 processed proteins & peptides Proteins 0.000 title claims abstract description 25
- 238000000034 method Methods 0.000 title claims abstract description 9
- 230000006698 induction Effects 0.000 title description 2
- 108090000623 proteins and genes Proteins 0.000 claims abstract description 9
- 102000004169 proteins and genes Human genes 0.000 claims abstract description 9
- 239000002671 adjuvant Substances 0.000 claims abstract description 6
- 210000004408 hybridoma Anatomy 0.000 claims abstract description 6
- 241001465754 Metazoa Species 0.000 claims abstract description 5
- 230000007910 cell fusion Effects 0.000 claims abstract description 5
- 102000014914 Carrier Proteins Human genes 0.000 claims abstract description 3
- 108010078791 Carrier Proteins Proteins 0.000 claims abstract description 3
- 108010008281 Recombinant Fusion Proteins Proteins 0.000 claims abstract description 3
- 230000008878 coupling Effects 0.000 claims 1
- 238000010168 coupling process Methods 0.000 claims 1
- 238000005859 coupling reaction Methods 0.000 claims 1
- 230000036039 immunity Effects 0.000 claims 1
- 230000001939 inductive effect Effects 0.000 abstract description 3
- 238000004519 manufacturing process Methods 0.000 description 2
- 241000699670 Mus sp. Species 0.000 description 1
- 102000007056 Recombinant Fusion Proteins Human genes 0.000 description 1
- 238000003745 diagnosis Methods 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000002649 immunization Methods 0.000 description 1
- 230000003053 immunization Effects 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 230000002194 synthesizing effect Effects 0.000 description 1
Landscapes
- Peptides Or Proteins (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
本发明属于生物工程技术领域，涉及用表位多肽诱导/制备表位特异性的单克隆抗体的方法。本发明首先人工合成带有天然蛋白或基因重组蛋白上的特定表位的表位多肽；其一个或多个表位经常是以重复的形式出现在人工合成的多肽(即表位多肽)上；其表位多肽耦联到载体蛋白或载体多肽上；其表位多肽的耦联物配以适合的佐剂免疫动物；采用常规的细胞融合技术制备杂交瘤。本发明能够在没有天然蛋白或基因重组的蛋白的条件下，高效地直接诱导/制备出预先确定的表位特异性单克隆抗体。The invention belongs to the technical field of bioengineering, and relates to a method for inducing/preparing epitope-specific monoclonal antibodies with epitope polypeptides. The present invention first artificially synthesizes epitope polypeptides with specific epitopes on natural proteins or gene recombinant proteins; one or more epitopes often appear on artificially synthesized polypeptides (i.e. epitope polypeptides) in repeated forms; The epitope polypeptide is coupled to the carrier protein or the carrier polypeptide; the epitope polypeptide conjugate is matched with a suitable adjuvant to immunize animals; the conventional cell fusion technique is used to prepare hybridomas. The present invention can directly induce/prepare the predetermined epitope-specific monoclonal antibody efficiently and efficiently without natural protein or genetically recombined protein.
Description
技术领域 本发明属于生物工程技术领域，特别涉及表位特异性的单克隆抗体制备及生产。Technical Field The present invention belongs to the technical field of bioengineering, in particular to the preparation and production of epitope-specific monoclonal antibodies.
背景技术 单克隆抗体技术(杂交瘤技术)是1975年发明的，1984年获得诺贝尔奖。该技术对生命科学和医学研究产生了划时代的影响，使疾病的诊断和治疗技术产生了革命性变化，并且造就了全球每年百亿美元的产业。目前通行的单克隆抗体技术是利用天然蛋白或基因重组的蛋白免疫动物，然后通过细胞融合技术制备分泌单克隆抗体的杂交瘤克隆。Background technology Monoclonal antibody technology (hybridoma technology) was invented in 1975 and won the Nobel Prize in 1984. This technology has had an epoch-making impact on life science and medical research, revolutionized the diagnosis and treatment of diseases, and created a global industry worth tens of billions of dollars per year. The current monoclonal antibody technology is to immunize animals with natural proteins or genetically recombined proteins, and then prepare hybridoma clones that secrete monoclonal antibodies through cell fusion technology.
现行单克隆抗体技术有无法克服的两个难题：其一，天然蛋白或基因重组的蛋白无法得到或很难提取，因而不能制备单克隆抗体；其二，往往要经过大量筛选才能得到分离得到需要的表位特异性的单克隆抗体。There are two insurmountable problems in the current monoclonal antibody technology: first, the natural protein or genetically recombined protein cannot be obtained or is difficult to extract, so the monoclonal antibody cannot be prepared; epitope-specific monoclonal antibodies.
发明内容 本发明的目的是为解决已有技术存在的难题，提出用表位多肽诱导/制备表位特异性的单克隆抗体的方法，能够在没有天然蛋白或基因重组的蛋白的条件下，高效地直接诱导/制备出预先确定的表位特异性单克隆抗体。SUMMARY OF THE INVENTION The purpose of the present invention is to solve the problems existing in the prior art, and propose a method for inducing/preparing epitope-specific monoclonal antibodies with epitope polypeptides, which can efficiently Direct induction/production of predetermined epitope-specific monoclonal antibodies.
本发明提出的用表位多肽诱导/制备表位特异性的单克隆抗体的方法，其特征在于包括以下步骤：The method for inducing/preparing epitope-specific monoclonal antibodies with epitope polypeptides proposed by the present invention is characterized in that it comprises the following steps:
1)人工合成带有天然蛋白或基因重组蛋白上的重复表位多肽C-(GPGRAFY)4；1) Artificially synthesized polypeptide C-(GPGRAFY) 4 with repeated epitopes on natural proteins or recombinant proteins;
2)所说的表位经常是以重复的形式出现在人工合成的多肽(即表位多肽)上；2) The said epitope often appears on the artificially synthesized polypeptide (i.e. epitope polypeptide) in repeated form;
3)所说的表位多肽耦联到载体蛋白或载体多肽上；3) The epitope polypeptide is coupled to a carrier protein or carrier polypeptide;
4)所说的表位多肽的耦联物配以适合佐剂免疫动物；4) The conjugate of the epitope polypeptide is matched with a suitable adjuvant to immunize animals;
5)采用常规的细胞融合技术制备杂交瘤。5) Hybridomas were prepared using conventional cell fusion techniques.
发明的效果：我们已经使用该项技术成功地制备了两株表位特异性的单克隆抗体。Effects of the invention: We have successfully prepared two epitope-specific monoclonal antibodies using this technique.
具体实施方式Detailed ways
实施例：制备GPGRAFY特异性的单克隆抗体Example: Preparation of GPGRAFY-specific monoclonal antibodies
步骤一，合成重复表位多肽C-(GPGRAFY)4；Step 1, synthesizing repeat epitope polypeptide C-(GPGRAFY) 4 ;
步骤二，利用MBS(m-maleimidoberrzoyl-N-hydroxy succinimide ester)将表位多肽与载体蛋白BSA耦联；Step 2, using MBS (m-maleimidoberrzoyl-N-hydroxy succinimide ester) to couple the epitope polypeptide to the carrier protein BSA;
步骤三，将耦联载体的表位多肽与福氏佐剂混合免疫Balb/c雌性小鼠，第一次用完全福氏佐剂，以后用不完全福氏佐剂。免疫剂量为10μg/每次/每只；每两周免疫一次；Step 3, the epitope polypeptide coupled with the carrier is mixed with Freund's adjuvant to immunize Balb/c female mice, the complete Freund's adjuvant is used for the first time, and the incomplete Freund's adjuvant is used thereafter. The immunization dose is 10 μg/each time/animal; once every two weeks;
步骤四，细胞融合实验；Step 4, cell fusion experiment;
步骤五，杂交瘤筛选。Step five, hybridoma screening.
Claims (1)
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB991238087A CN1152053C (en) | 1999-11-12 | 1999-11-12 | Method for induction and prepn. of epitope idiosyncratic monoclonal antibody by using epitope polypeptide |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB991238087A CN1152053C (en) | 1999-11-12 | 1999-11-12 | Method for induction and prepn. of epitope idiosyncratic monoclonal antibody by using epitope polypeptide |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1259524A CN1259524A (en) | 2000-07-12 |
| CN1152053C true CN1152053C (en) | 2004-06-02 |
Family
ID=5283004
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CNB991238087A Expired - Fee Related CN1152053C (en) | 1999-11-12 | 1999-11-12 | Method for induction and prepn. of epitope idiosyncratic monoclonal antibody by using epitope polypeptide |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN1152053C (en) |
Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110790829B (en) * | 2017-12-19 | 2022-08-02 | 北京泽勤生物医药有限公司 | Application of antibody prepared by using pHLIP extracellular segment as antigen in preparation of antitumor drugs |
| CN110272502B (en) * | 2019-07-12 | 2021-09-14 | 深圳市亚辉龙生物科技股份有限公司 | Immunogen, hybridoma cell secreting anti-cardiac troponin I monoclonal antibody, preparation method, monoclonal antibody and application |
-
1999
- 1999-11-12 CN CNB991238087A patent/CN1152053C/en not_active Expired - Fee Related
Also Published As
| Publication number | Publication date |
|---|---|
| CN1259524A (en) | 2000-07-12 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Schroer et al. | Mapping epitopes on the insulin molecule using monoclonal antibodies | |
| US8815244B2 (en) | Method for production of antibody using ostrich | |
| US4879225A (en) | Enhanced production of antibodies utilizing insolubilized immune complexes | |
| CN112961222A (en) | 2019 novel coronavirus N protein linear epitope peptide, monoclonal antibody and application | |
| ATE225668T1 (en) | NEW METHOD OF VACCINATION AND VACCINES THEREOF CONTAINING AN EPITOPE-CODING NUCLEIC ACID AND AN EPITOPE-CONTAINING PEPTIDE | |
| JPH02488A (en) | Antihuman bcdf monmoclonal antibody | |
| JPS62294697A (en) | Synthetic peptide and immunogenic composition | |
| CN106866820B (en) | Monoclonal antibody for capturing tumor cells and resisting human keratin 18 and application thereof | |
| AU702294B2 (en) | Proteins from mammalian liver and their use in oncology | |
| JPS60155133A (en) | Anti-idio type vaccine using anti-idio type monoclonal antibody | |
| WO2015165368A1 (en) | Anti-her2 neutralizing activity monoclonal antibody and use thereof | |
| CN1152053C (en) | Method for induction and prepn. of epitope idiosyncratic monoclonal antibody by using epitope polypeptide | |
| US5084396A (en) | Enhanced production of antibodies utilizing insolubilized immune complexes | |
| JPH02503383A (en) | human tumor-associated antigens | |
| CN113061177A (en) | CD73 enzyme activity related antigen epitope and preparation method of specific antibody aiming at epitope | |
| Meyers et al. | Human antiglomerular basement membrane autoantibody disease in XenoMouse II | |
| CN104610441B (en) | For preparing artificial semiantigen, preparation method and the monoclonal antibody of acquisition of glypican-3 (GPC3) monoclonal antibody | |
| JPH06500992A (en) | Immunogenic peptide sequence of Echinococcus granulosus, DNA sequence encoding this peptide sequence, and diagnostic and therapeutic uses | |
| CN109679924A (en) | The anti-human monoclonal antibody and the preparation method and application thereof with peptide element of high-affinity | |
| CN106117341B (en) | Antigen epitope peptide of IL-9 and its application | |
| Domingue et al. | Antibodies to bacterial vaccines demonstrating specificity for human choriogonadotropin (hCG) and immunochemical detection of hCG-like factor in subcellular bacterial fractions | |
| CN110054663B (en) | CKAP4 epitope peptide, antigen, monoclonal antibody, application of monoclonal antibody and CKAP4 detection test strip | |
| Majidi et al. | Production and characterization of monoclonal antibodies against human IgG in Balb/c mouse | |
| LT5414B (en) | Process for preparing monoclonal antibodies; hybridoma cell lines generated by this process and recombinant chimeric virus-like particles with inserted foreign protein fragments as immunogenes to generate hybridomas by this process | |
| JP2994074B2 (en) | Determination of cedar and cypress pollen |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C06 | Publication | ||
| PB01 | Publication | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| REG | Reference to a national code |
Ref country code: HK Ref legal event code: GR Ref document number: 1066262 Country of ref document: HK |
|
| C19 | Lapse of patent right due to non-payment of the annual fee | ||
| CF01 | Termination of patent right due to non-payment of annual fee |