JPH0787799B2 - Monoclonal antibody production enhancement method - Google Patents
Monoclonal antibody production enhancement methodInfo
- Publication number
- JPH0787799B2 JPH0787799B2 JP63077043A JP7704388A JPH0787799B2 JP H0787799 B2 JPH0787799 B2 JP H0787799B2 JP 63077043 A JP63077043 A JP 63077043A JP 7704388 A JP7704388 A JP 7704388A JP H0787799 B2 JPH0787799 B2 JP H0787799B2
- Authority
- JP
- Japan
- Prior art keywords
- medium
- antibody
- monoclonal antibody
- human
- antibody production
- 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
Landscapes
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Description
【発明の詳細な説明】 産業上の利用分野 本発明は、抗体生産細胞によるモノクローナル抗体の生
産を増強する方法に関するものである。TECHNICAL FIELD The present invention relates to a method for enhancing the production of monoclonal antibodies by antibody-producing cells.
従来の技術及び発明が解決しようとする課題 従来抗体生産細胞をペトリ皿、又は培養槽などを用いて
培養し抗体を生産する場合、生産効率が悪く大量に入手
することが困難であった。特にヒト−ヒトハイブリドー
マを用いてヒト型モノクローナル抗体を生産しようとす
る場合には、生産量が極端に少なく、その利用に限界が
あった。2. Description of the Related Art When producing antibody by culturing antibody-producing cells in a conventional Petri dish or a culture tank, production efficiency is poor and it is difficult to obtain a large amount. Particularly, in the case of producing a human type monoclonal antibody using a human-human hybridoma, the production amount was extremely small, and its utilization was limited.
この発明は、このような問題点を解決しようとして行わ
れたものである。The present invention has been made in order to solve such a problem.
課題を解決するための手段 本発明者らは、モノクローナル抗体の生産を上昇させる
培養方法の検討を行った結果、抗体生産細胞の増殖速度
又は生存率を調節することがこの目的に適合することを
見いだし、これに基づいて本発明を完成するに至つた。Means for Solving the Problems As a result of investigating a culture method for increasing the production of monoclonal antibodies, the present inventors have found that controlling the growth rate or survival rate of antibody-producing cells is suitable for this purpose. The present invention has been found and based on this, the present invention has been completed.
以下に本発明について詳細に説明する。The present invention will be described in detail below.
（１）抗体生産細胞 ヒト−ヒトハイブリドーマ、マウス−マウスハイブリド
ーマ、EBウィルスで形質転換した細胞等が利用できる。
特に、ヒト−ヒトハイブリドーマに有効である。(1) Antibody-producing cells Human-human hybridomas, mouse-mouse hybridomas, cells transformed with EB virus, and the like can be used.
In particular, it is effective for human-human hybridomas.
（２）細胞培養に用いられる培地 通常用いられる無血清培地又は血清培地が利用できる。
血清培地としては10％牛胎児血清を添加した培地が例示
できるが、モノクローナル抗体生産用培地としては、抗
体の精製が容易であること、培地が安価であること等の
理由から無血清培地が望ましい。無血清培地としては、
基本合成培地に血清アルブミン、インシュリン、トラン
スフェリン、エタノールアミン、セレニウム等の因子を
添加した培地が用いられる。基本合成培地としてはダル
ベッコMEM培地（DME）、ハムF-12培地、RDF培地（PRMI
1640、DME及びハムＦ−12培地を2:1:1で混合した培地）
等が用いられるが、好ましくはRDF培地が用いられる。(2) Medium used for cell culture A commonly used serum-free medium or serum medium can be used.
As the serum medium, a medium supplemented with 10% fetal bovine serum can be exemplified, but as the medium for producing the monoclonal antibody, a serum-free medium is preferable because of easy purification of the antibody, inexpensive medium and the like. . As a serum-free medium,
A medium in which factors such as serum albumin, insulin, transferrin, ethanolamine, and selenium are added to the basic synthetic medium is used. Dulbecco's MEM medium (DME), Ham's F-12 medium, RDF medium (PRMI)
1640, DME and Ham F-12 medium mixed at 2: 1: 1)
Etc. are used, but RDF medium is preferably used.
（３）培養法 ペトリ皿、灌流型連続培養装置、又はホローファイバー
型の培養装置が利用できる。(3) Culture method A Petri dish, a perfusion type continuous culture apparatus, or a hollow fiber type culture apparatus can be used.
（４）抗体生産細胞の増殖速度及び生存率の調節 増殖速度及び生存率を調節する方法としては、25−60mM
塩化カリウムを（２）に示した培地に添加する方法、灌
流型培養装置で培養する際に灌流速度を調節する方法、
培養温度を低下又は上昇させる方法等がある。好ましく
は30mM塩化カリウムの添加により行う。(4) Regulation of proliferation rate and survival rate of antibody-producing cells As a method for regulating the proliferation rate and survival rate, 25-60 mM
A method of adding potassium chloride to the medium shown in (2), a method of adjusting the perfusion rate when culturing in a perfusion type culture device,
There is a method of lowering or raising the culture temperature. Preference is given to adding 30 mM potassium chloride.
発明の効果 塩化カリウムを培地へ添加して、抗体生産細胞の増殖速
度及び生存率を低下させることにより、抗体生産量が1.
5倍に上昇した。このことから、モノクローナル抗体の
生産には、細胞が旺盛に増殖している状態より、細胞の
増殖速度が低下し生存率の低下しつつある状態の方が有
利と考えられる。EFFECTS OF THE INVENTION By adding potassium chloride to a medium to reduce the growth rate and viability of antibody-producing cells, the amount of antibody produced is 1.
It rose five times. From this, it is considered that the production of the monoclonal antibody is more advantageous in the state in which the cell growth rate is decreasing and the survival rate is decreasing than in the state in which the cells are actively growing.
以下本発明の実施例を示す。Examples of the present invention will be shown below.
実施例１ IgM型抗体生産性ヒト−ヒトハイブリドーマHB4C5〔イン
・ビトロ・セルラー・アンド・ディベロップメンタル・
バイオロジー（In Vitoro Cell.Develop.Biol.）、21、
593（1985）〕を、35mmプラスチックペトリ皿に5x105細
胞/mlになるように下記の培地中に植え込んだ。Example 1 IgM type antibody-producing human-human hybridoma HB4C5 [in vitro cellular and developmental
Biology (In Vitoro Cell.Develop.Biol.), 21 ,
593 (1985)] was plated in a 35 mm plastic petri dish at 5 × 10 5 cells / ml in the following medium.
非添加培地 RDFに下記のものを添加する。Non-supplemented medium Add the following to RDF.
血清アルブミン （ヒト） 2mg/ml インシュリン （ウシ） 5μg/ml トランスフェリン（ヒト） 35μg/ml エタノールアミン 20μＭ セレニウム 2.5nM 塩化カリウム添加培地 に塩化カリウムを30mMになるように添加する。Serum albumin (human) 2 mg / ml Insulin (bovine) 5 μg / ml Transferrin (human) 35 μg / ml Ethanolamine 20 μM Selenium 2.5 nM Potassium chloride is added to the medium to 30 mM potassium chloride.
２日間培養して抗体濃度、細胞密度及び生存率を測定し
たとこ、第１表に示したように非添加培地に比べ塩化カ
リウム添加培地は細胞増殖速度及び生存率を低下させる
傾向が認められ、抗体生産を1.5倍に増強した。After culturing for 2 days and measuring the antibody concentration, cell density and viability, as shown in Table 1, potassium chloride-added medium was observed to tend to lower the cell growth rate and viability as compared with the non-added medium. Antibody production was increased 1.5-fold.
Claims (1)
となるように添加することを特徴とするモノクローナル
抗体生産増強法。1. The final concentration of potassium chloride in the medium is 25-60 mM.
A method for enhancing the production of a monoclonal antibody, characterized in that
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP63077043A JPH0787799B2 (en) | 1988-03-30 | 1988-03-30 | Monoclonal antibody production enhancement method |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP63077043A JPH0787799B2 (en) | 1988-03-30 | 1988-03-30 | Monoclonal antibody production enhancement method |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPH01247097A JPH01247097A (en) | 1989-10-02 |
| JPH0787799B2 true JPH0787799B2 (en) | 1995-09-27 |
Family
ID=13622748
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP63077043A Expired - Fee Related JPH0787799B2 (en) | 1988-03-30 | 1988-03-30 | Monoclonal antibody production enhancement method |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH0787799B2 (en) |
Families Citing this family (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| SG187397A1 (en) * | 2007-12-27 | 2013-02-28 | Baxter Int | Cell culture processes |
| SG11201909087PA (en) | 2017-03-31 | 2019-10-30 | Boehringer Ingelheim Int | Perfusion medium |
| JP7513034B2 (en) | 2019-11-05 | 2024-07-09 | 味の素株式会社 | Protein Production Methods |
Family Cites Families (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPS6251983A (en) * | 1985-09-02 | 1987-03-06 | Hagiwara Yoshihide | Serum-free culture medium for cultivating human/human hybridoma |
| JPS62269694A (en) * | 1986-05-20 | 1987-11-24 | Teijin Ltd | Method for cultivating multiplicative animal cell to produce useful high polymer substance and culture system therefor |
| FR2600076A1 (en) * | 1986-06-12 | 1987-12-18 | Fond Ctre Nal Transfusion | CULTURE MEDIUM COMPRISING HUMAN ALBUMIN, PROCESS FOR PREPARING A PRODUCT INJECTABLE THEREFROM, PRODUCT OBTAINED AND USE THEREOF, COMPOSITION OBTAINED |
| JPH084518B2 (en) * | 1986-08-30 | 1996-01-24 | 萩原 義秀 | Serum-free culture method for human / human hybridoma producing monoclonal antibody |
-
1988
- 1988-03-30 JP JP63077043A patent/JPH0787799B2/en not_active Expired - Fee Related
Also Published As
| Publication number | Publication date |
|---|---|
| JPH01247097A (en) | 1989-10-02 |
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