JPH0543349B2 - - Google Patents
Info
- Publication number
- JPH0543349B2 JPH0543349B2 JP58050184A JP5018483A JPH0543349B2 JP H0543349 B2 JPH0543349 B2 JP H0543349B2 JP 58050184 A JP58050184 A JP 58050184A JP 5018483 A JP5018483 A JP 5018483A JP H0543349 B2 JPH0543349 B2 JP H0543349B2
- Authority
- JP
- Japan
- Prior art keywords
- fraction
- medium
- molecular weight
- cells
- royal jelly
- 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 - Lifetime
Links
- 210000004369 blood Anatomy 0.000 claims description 15
- 239000008280 blood Substances 0.000 claims description 15
- 230000003169 placental effect Effects 0.000 claims description 14
- 239000006143 cell culture medium Substances 0.000 claims description 3
- 229940109850 royal jelly Drugs 0.000 description 13
- 239000002609 medium Substances 0.000 description 12
- 210000004027 cell Anatomy 0.000 description 10
- 235000018102 proteins Nutrition 0.000 description 10
- 102000004169 proteins and genes Human genes 0.000 description 10
- 108090000623 proteins and genes Proteins 0.000 description 10
- 235000013336 milk Nutrition 0.000 description 9
- 239000008267 milk Substances 0.000 description 9
- 210000004080 milk Anatomy 0.000 description 9
- 210000004102 animal cell Anatomy 0.000 description 8
- 238000012258 culturing Methods 0.000 description 7
- 239000001963 growth medium Substances 0.000 description 7
- 210000002966 serum Anatomy 0.000 description 7
- 239000000126 substance Substances 0.000 description 7
- 102000010445 Lactoferrin Human genes 0.000 description 4
- 108010063045 Lactoferrin Proteins 0.000 description 4
- 239000007640 basal medium Substances 0.000 description 4
- NOESYZHRGYRDHS-UHFFFAOYSA-N insulin Chemical compound N1C(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(NC(=O)CN)C(C)CC)CSSCC(C(NC(CO)C(=O)NC(CC(C)C)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CCC(N)=O)C(=O)NC(CC(C)C)C(=O)NC(CCC(O)=O)C(=O)NC(CC(N)=O)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CSSCC(NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2C=CC(O)=CC=2)NC(=O)C(CC(C)C)NC(=O)C(C)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2NC=NC=2)NC(=O)C(CO)NC(=O)CNC2=O)C(=O)NCC(=O)NC(CCC(O)=O)C(=O)NC(CCCNC(N)=N)C(=O)NCC(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC(O)=CC=3)C(=O)NC(C(C)O)C(=O)N3C(CCC3)C(=O)NC(CCCCN)C(=O)NC(C)C(O)=O)C(=O)NC(CC(N)=O)C(O)=O)=O)NC(=O)C(C(C)CC)NC(=O)C(CO)NC(=O)C(C(C)O)NC(=O)C1CSSCC2NC(=O)C(CC(C)C)NC(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(N)CC=1C=CC=CC=1)C(C)C)CC1=CN=CN1 NOESYZHRGYRDHS-UHFFFAOYSA-N 0.000 description 4
- CSSYQJWUGATIHM-IKGCZBKSSA-N l-phenylalanyl-l-lysyl-l-cysteinyl-l-arginyl-l-arginyl-l-tryptophyl-l-glutaminyl-l-tryptophyl-l-arginyl-l-methionyl-l-lysyl-l-lysyl-l-leucylglycyl-l-alanyl-l-prolyl-l-seryl-l-isoleucyl-l-threonyl-l-cysteinyl-l-valyl-l-arginyl-l-arginyl-l-alanyl-l-phenylal Chemical compound C([C@H](N)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CS)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(=O)NCC(=O)N[C@@H](C)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CO)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CS)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](C)C(=O)N[C@@H](CC=1C=CC=CC=1)C(O)=O)C1=CC=CC=C1 CSSYQJWUGATIHM-IKGCZBKSSA-N 0.000 description 4
- 229940078795 lactoferrin Drugs 0.000 description 4
- 235000021242 lactoferrin Nutrition 0.000 description 4
- 210000004698 lymphocyte Anatomy 0.000 description 3
- 239000000243 solution Substances 0.000 description 3
- 241000282412 Homo Species 0.000 description 2
- 102000004877 Insulin Human genes 0.000 description 2
- 108090001061 Insulin Proteins 0.000 description 2
- IQFYYKKMVGJFEH-XLPZGREQSA-N Thymidine Chemical compound O=C1NC(=O)C(C)=CN1[C@@H]1O[C@H](CO)[C@@H](O)C1 IQFYYKKMVGJFEH-XLPZGREQSA-N 0.000 description 2
- 239000000654 additive Substances 0.000 description 2
- 239000000872 buffer Substances 0.000 description 2
- 238000000502 dialysis Methods 0.000 description 2
- LOKCTEFSRHRXRJ-UHFFFAOYSA-I dipotassium trisodium dihydrogen phosphate hydrogen phosphate dichloride Chemical compound P(=O)(O)(O)[O-].[K+].P(=O)(O)([O-])[O-].[Na+].[Na+].[Cl-].[K+].[Cl-].[Na+] LOKCTEFSRHRXRJ-UHFFFAOYSA-I 0.000 description 2
- 238000011067 equilibration Methods 0.000 description 2
- 239000012847 fine chemical Substances 0.000 description 2
- 238000005194 fractionation Methods 0.000 description 2
- 238000002523 gelfiltration Methods 0.000 description 2
- 235000020256 human milk Nutrition 0.000 description 2
- 210000004251 human milk Anatomy 0.000 description 2
- FDGQSTZJBFJUBT-UHFFFAOYSA-N hypoxanthine Chemical compound O=C1NC=NC2=C1NC=N2 FDGQSTZJBFJUBT-UHFFFAOYSA-N 0.000 description 2
- 229940125396 insulin Drugs 0.000 description 2
- 238000000034 method Methods 0.000 description 2
- 239000002953 phosphate buffered saline Substances 0.000 description 2
- 210000002826 placenta Anatomy 0.000 description 2
- 239000006228 supernatant Substances 0.000 description 2
- NHBKXEKEPDILRR-UHFFFAOYSA-N 2,3-bis(butanoylsulfanyl)propyl butanoate Chemical compound CCCC(=O)OCC(SC(=O)CCC)CSC(=O)CCC NHBKXEKEPDILRR-UHFFFAOYSA-N 0.000 description 1
- QKNYBSVHEMOAJP-UHFFFAOYSA-N 2-amino-2-(hydroxymethyl)propane-1,3-diol;hydron;chloride Chemical compound Cl.OCC(N)(CO)CO QKNYBSVHEMOAJP-UHFFFAOYSA-N 0.000 description 1
- DWRXFEITVBNRMK-UHFFFAOYSA-N Beta-D-1-Arabinofuranosylthymine Natural products O=C1NC(=O)C(C)=CN1C1C(O)C(O)C(CO)O1 DWRXFEITVBNRMK-UHFFFAOYSA-N 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 1
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 101000798114 Homo sapiens Lactotransferrin Proteins 0.000 description 1
- UGQMRVRMYYASKQ-UHFFFAOYSA-N Hypoxanthine nucleoside Natural products OC1C(O)C(CO)OC1N1C(NC=NC2=O)=C2N=C1 UGQMRVRMYYASKQ-UHFFFAOYSA-N 0.000 description 1
- 102000014150 Interferons Human genes 0.000 description 1
- 108010050904 Interferons Proteins 0.000 description 1
- VCUFZILGIRCDQQ-KRWDZBQOSA-N N-[[(5S)-2-oxo-3-(2-oxo-3H-1,3-benzoxazol-6-yl)-1,3-oxazolidin-5-yl]methyl]-2-[[3-(trifluoromethoxy)phenyl]methylamino]pyrimidine-5-carboxamide Chemical compound O=C1O[C@H](CN1C1=CC2=C(NC(O2)=O)C=C1)CNC(=O)C=1C=NC(=NC=1)NCC1=CC(=CC=C1)OC(F)(F)F VCUFZILGIRCDQQ-KRWDZBQOSA-N 0.000 description 1
- 206010035226 Plasma cell myeloma Diseases 0.000 description 1
- 239000012980 RPMI-1640 medium Substances 0.000 description 1
- BUGBHKTXTAQXES-UHFFFAOYSA-N Selenium Chemical compound [Se] BUGBHKTXTAQXES-UHFFFAOYSA-N 0.000 description 1
- 229920002684 Sepharose Polymers 0.000 description 1
- 108010071390 Serum Albumin Proteins 0.000 description 1
- 102000007562 Serum Albumin Human genes 0.000 description 1
- 102000004338 Transferrin Human genes 0.000 description 1
- 108090000901 Transferrin Proteins 0.000 description 1
- IQFYYKKMVGJFEH-UHFFFAOYSA-N beta-L-thymidine Natural products O=C1NC(=O)C(C)=CN1C1OC(CO)C(O)C1 IQFYYKKMVGJFEH-UHFFFAOYSA-N 0.000 description 1
- 244000309466 calf Species 0.000 description 1
- 238000004113 cell culture Methods 0.000 description 1
- 230000010261 cell growth Effects 0.000 description 1
- 230000004663 cell proliferation Effects 0.000 description 1
- 238000010828 elution Methods 0.000 description 1
- 229940088598 enzyme Drugs 0.000 description 1
- 239000012091 fetal bovine serum Substances 0.000 description 1
- 210000002950 fibroblast Anatomy 0.000 description 1
- 230000004927 fusion Effects 0.000 description 1
- 230000012010 growth Effects 0.000 description 1
- 102000050459 human LTF Human genes 0.000 description 1
- 210000005260 human cell Anatomy 0.000 description 1
- 210000004408 hybridoma Anatomy 0.000 description 1
- 229940079322 interferon Drugs 0.000 description 1
- 244000144972 livestock Species 0.000 description 1
- 201000000050 myeloid neoplasm Diseases 0.000 description 1
- 102000039446 nucleic acids Human genes 0.000 description 1
- 108020004707 nucleic acids Proteins 0.000 description 1
- 150000007523 nucleic acids Chemical class 0.000 description 1
- 230000002093 peripheral effect Effects 0.000 description 1
- 239000002243 precursor Substances 0.000 description 1
- 229910052711 selenium Inorganic materials 0.000 description 1
- 239000011669 selenium Substances 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 229940104230 thymidine Drugs 0.000 description 1
- 239000012581 transferrin Substances 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
Landscapes
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Description
本発明は、動物細胞または動物細胞由来の細胞
を培養する低タンパク質濃度の培地に関するもの
であり、胎盤血の分子量5−8万の画分を含有す
ることを特徴とする細胞培養用培地に関するもの
である。
動物細胞および動物細胞由来の細胞には、抗
体、インターフエロン、インシユリン、酵素など
の有効物質を産生するものが知られており、その
細胞を培養することによつてこれらの有効物質を
得ることの実施または試みがなされている。従来
よりこれらの細胞を培養するためには、ほとんど
の場合、培地に血清を添加することが必要であ
る。血清としては一般に牛胎児血清、小牛血清な
どが用いられており、添加量は通常10%程度であ
る。しかし、これらの血清は高価であること、品
質が一定しないこと、供給に制限があることなど
の難点がある。
本発明者らは、これらのことを解決すべく研究
を重ねた結果、低タンパク質濃度の培地で細胞増
殖が可能なことを見いだし、本発明を完成したも
のである。
胎盤はヒト、家畜(牛、馬など)からのもので
ある。胎盤血の分子量5−8万の画分(以下胎盤
血画分と略す)は、例えば胎盤から自然にしみ出
た血液を2万×gで遠心し、その上清を更にSe
−phadex G−200(Pharmacia Fine
Chemicals)、Cellulofine GC−700(生化学工業
株式会社)などを用いるゲルろ過法により分子量
分画して得ることができる。そのゲルろ過に用い
る平衡化および溶出用溶液は、例えばリン酸緩衝
化生理食塩水、100mM Na−Hepes緩衝液(PH
7.4)、水などである。本発明の細胞培養用培地
は、この胎盤血画分を通常の動物細胞の培養に用
いられる基礎培地(血清を含まない)に添加して
用いる。
また、これに乳および/または乳の成分、ロー
ヤルゼリーおよび/またはローヤルゼリーの成分
の中より1種以上を基礎培地に添加して用いるこ
ともできる。
乳の成分は、以下のようにして得る。乳を例え
ば2万×gで遠心して溶液画分(以下乳の遠心溶
液画分と略す)を得る。乳の分子量20万以上の画
分および分子量4−9万の画分は、胎盤血の分子
量分画と同様の方法で得ることができる。ラクト
フエリンは例えばLo¨nnerdalらの方法
(Lo¨nnerdal、B.、Carlsson、J.、and Porath、
J(1977)FEBS Lett.75、89−92)で精製する。
また、ローヤルゼリーの成分は、以下のように
して得る。ローヤルゼリーの分子量20万以上の画
分および分子量5−8万の画分は、胎盤血の分子
量分画と同様の方法で得ることができる。ローヤ
ルゼリーの分子量約58000のタンパク質(以下ロ
ーヤルゼリータンパク質と略す)は、例えば以下
のようにして精製する。ローヤルゼリーをリン酸
緩衝化生理食塩水で2倍に希釈後、それに対し透
析する。透析後、2万×gで遠心し、得られる上
清を胎盤血の場合と同様の方法で分子量分画し、
5−8万の分子量画分を得る。この画分を1mM
Tris−HCl緩衝液(PH8.0)に透析後、同様の
緩衝液で平衡化しBlue Sepharose(Pharmacia
Fine Chemicals)にかけ、素通り画分すなわち
ローヤルゼリータンパク質を得る。
本発明培地の基礎培地としては、一般の動物細
胞の培養に用いられるものであればいかなるもの
も用いることができる。例えば、Dulbecco′s
Modified Eagle medium(以下D′MEMと略す)、
F−12、RPMI−1640などの基礎培地、また、こ
れら基礎培地にインシユリン(2−10μg/ml)、
セレニウム(10-11−10-8M)、トランスフエリン
(10−35μg/ml)、核酸前駆体(例えばチミジ
ン、ヒポキサンチンなど、10-6M程度)、血清ア
ルブミン(50−400μg/ml)などを添加した培
地も基礎培地として用いることができる。本発明
の培地に添加する胎盤血画分の最終タンパク質濃
度は50−300μg/mlであることが望ましい。そ
の他の乳および乳の成分については0.5−350μ
g/ml、ローヤルゼリーおよびローヤルゼリーの
成分については10−200μg/mlが望ましい。
本発明培地は、繊維芽細胞、リンパ芽球細胞な
どの動物細胞の培養に用いることもできるが、リ
ンパ球由来細胞などの動物細胞由来の細胞の培養
に最も好適に用いられる。本発明の培地を用いれ
ば血清を大量に添加した従来の培地にほぼ匹敵す
る細胞増殖を得ることができ、しかも従来の培地
と比べコストダウンが期待できる。また、血清を
10%添加した場合の培地中のタンパク質濃度が3
mg/ml以上であるのに比べ、本発明培地は0.4
mg/ml以下と低タンパク質濃度であることから、
有効物質産生に本発明培地を用いることにより有
効物質の分離・精製が容易となる。更に、本発明
培地は分子量が5−8万の画分を添加することか
ら、有効物質の分子量が5−8万の範囲外の場合
には、有効物質の分離・精製が容易となる。例え
ば抗体(分子量15万以上)を産生する細胞を培養
する場合には、胎盤血画分と、ラクトフエリンお
よび/または乳の分子量4−9万の画分、ローヤ
ルゼリータンパク質および/またはローヤルゼリ
ーの分子量5−8万の画分を必要に応じ加え、用
いるなどのように行う。更に、ヒトの胎盤血画分
と、要すればヒトラクトフエリンなどの人乳成分
および/または人乳を添加した培地でヒトの細胞
を培養する場合には、培地中の成分がすべてヒト
由来であることから、培地中に産生した有効物質
を完全に精製しなくともヒトに投与可能であると
いう利点を有する。
次に実施例により本発明を説明する。
実施例 1
健康人の末梢リンパ球株細胞(Bir7)の105個
細胞を、D′MEM/F−12(1:1)に第1表に
示す添加物を添加した培地2mlに、植え込む。5
%CO2、37℃の条件下で3日間培養を行う。得ら
れた結果は第1表に示すとおりである。
The present invention relates to a medium with a low protein concentration for culturing animal cells or cells derived from animal cells, and relates to a cell culture medium characterized by containing a fraction of placental blood with a molecular weight of 50,000 to 80,000. It is. Animal cells and cells derived from animal cells are known to produce effective substances such as antibodies, interferon, insulin, and enzymes, and it is possible to obtain these effective substances by culturing these cells. being carried out or attempted. Traditionally, in order to culture these cells, it is almost always necessary to add serum to the culture medium. Fetal bovine serum, calf serum, etc. are generally used as serum, and the amount added is usually about 10%. However, these serums have drawbacks such as being expensive, having inconsistent quality, and limited supply. As a result of repeated research to solve these problems, the present inventors discovered that cell growth is possible in a medium with a low protein concentration, and completed the present invention. Placentas can come from humans or livestock (cows, horses, etc.). Placental blood fraction with a molecular weight of 50,000 to 80,000 (hereinafter referred to as placental blood fraction) can be obtained by, for example, centrifuging blood naturally exuded from the placenta at 20,000 x g, and then subjecting the supernatant to further Separation.
-phadex G-200 (Pharmacia Fine
Chemicals), Cellulofine GC-700 (Seikagaku Corporation), etc. by gel filtration method to obtain molecular weight fractionation. Equilibration and elution solutions used for gel filtration include, for example, phosphate buffered saline, 100mM Na-Hepes buffer (PH
7.4), water, etc. The cell culture medium of the present invention is used by adding this placental blood fraction to a basal medium (not containing serum) used for normal animal cell culture. Furthermore, one or more of milk and/or milk components, royal jelly and/or royal jelly components may be added to the basal medium. The milk components are obtained as follows. Milk is centrifuged at, for example, 20,000×g to obtain a solution fraction (hereinafter referred to as milk centrifuged solution fraction). The fraction of milk with a molecular weight of 200,000 or more and the fraction with a molecular weight of 40,000 to 90,000 can be obtained in the same manner as the molecular weight fraction of placental blood. Lactoferrin can be obtained, for example, by the method of Lo¨nnerdal et al. (Lo¨nnerdal, B., Carlsson, J., and Porath,
J (1977) FEBS Lett. 75, 89-92). Moreover, the components of royal jelly can be obtained as follows. The fraction of royal jelly with a molecular weight of 200,000 or more and the fraction with a molecular weight of 50,000 to 80,000 can be obtained in the same manner as the molecular weight fraction of placental blood. Royal jelly protein having a molecular weight of about 58,000 (hereinafter abbreviated as royal jelly protein) is purified, for example, as follows. Royal jelly is diluted twice with phosphate buffered saline and then dialyzed against it. After dialysis, it was centrifuged at 20,000 x g, and the resulting supernatant was subjected to molecular weight fractionation in the same manner as for placental blood.
A molecular weight fraction of 50,000 to 80,000 is obtained. This fraction was added to 1mM
After dialysis against Tris-HCl buffer (PH8.0), equilibration with the same buffer and Blue Sepharose (Pharmacia
Fine Chemicals) to obtain a flow-through fraction, that is, royal jelly protein. As the basal medium of the culture medium of the present invention, any medium used for culturing general animal cells can be used. For example, Dulbecco's
Modified Eagle medium (hereinafter abbreviated as D′MEM),
Basal media such as F-12, RPMI-1640, insulin (2-10 μg/ml),
Selenium (10 -11 -10 -8 M), transferrin (10-35 μg/ml), nucleic acid precursors (e.g. thymidine, hypoxanthine, etc., about 10 -6 M), serum albumin (50-400 μg/ml), etc. A medium supplemented with can also be used as a basal medium. The final protein concentration of the placental blood fraction added to the culture medium of the present invention is preferably 50-300 μg/ml. 0.5−350μ for other milk and milk components
g/ml, 10-200 μg/ml for royal jelly and royal jelly components. Although the culture medium of the present invention can be used for culturing animal cells such as fibroblasts and lymphoblastoid cells, it is most preferably used for culturing animal cell-derived cells such as lymphocyte-derived cells. By using the medium of the present invention, it is possible to obtain cell proliferation that is almost comparable to that of a conventional medium containing a large amount of serum, and moreover, it is expected that the cost will be lower than that of the conventional medium. Also, serum
The protein concentration in the medium when adding 10% is 3
mg/ml or more, compared to 0.4 mg/ml for the culture medium of the present invention.
Due to its low protein concentration of less than mg/ml,
By using the culture medium of the present invention for producing an effective substance, it becomes easy to separate and purify the effective substance. Furthermore, since a fraction having a molecular weight of 50,000 to 80,000 is added to the culture medium of the present invention, if the molecular weight of the effective substance is outside the range of 50,000 to 80,000, the effective substance can be easily separated and purified. For example, when culturing cells that produce antibodies (molecular weight of 150,000 or more), placental blood fraction, lactoferrin and/or milk fraction with molecular weight of 40,000 to 90,000, royal jelly protein and/or royal jelly with molecular weight of 5 to 90,000 are used. 80,000 fractions are added and used as needed, and so on. Furthermore, when culturing human cells in a medium supplemented with a human placental blood fraction and, if necessary, human milk components such as human lactoferrin and/or human milk, it is necessary to ensure that all components in the medium are of human origin. Therefore, it has the advantage that the effective substance produced in the medium can be administered to humans without being completely purified. Next, the present invention will be explained with reference to examples. Example 1 10 5 cells of a peripheral lymphocyte cell line (Bir7) from a healthy person are implanted in 2 ml of a medium containing D'MEM/F-12 (1:1) and the additives shown in Table 1. 5
Culture is carried out under conditions of % CO 2 and 37° C. for 3 days. The results obtained are shown in Table 1.
【表】
なおD′MEM/F−12に添加物としてヒト胎盤
血画分単独、ヒト胎盤血画分およびラクトフエリ
ン、ヒト胎盤血画分とラクトフエリンおよびロー
ヤルゼリータンパク質の3種をそれぞれ加えた培
地で30日間以上継代培養してもそれぞれの増殖速
度に変化がみられなかつた。このことは永続的に
継代培養することが可能であることを示してい
る。
実施例 2
マウスのミエローマ(NS−1)とマウスB−
リンパ球(BALB/c)が融合して生成したハ
イブリドーマについて植え込みの細胞数を0.5×
105個にへらし、実施例1と同様にして培養を行
う。得られた結果は第2表に示すとおりである。[Table] In addition, D'MEM/F-12 was supplemented with three types of additives: human placental blood fraction alone, human placental blood fraction and lactoferrin, human placental blood fraction and lactoferrin, and royal jelly protein. No change was observed in the respective growth rates even after subculturing for more than one day. This indicates that continuous subculture is possible. Example 2 Mouse myeloma (NS-1) and mouse B-
For hybridomas generated by fusion of lymphocytes (BALB/c), the number of implanted cells is 0.5
Cut into 10 5 pieces and culture in the same manner as in Example 1. The results obtained are shown in Table 2.
【表】
%
【table】 %
Claims (1)
を特徴とする細胞培養用培地。1. A cell culture medium characterized by containing a fraction of placental blood with a molecular weight of 50,000 to 80,000.
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP58050184A JPS59175876A (en) | 1983-03-25 | 1983-03-25 | Medium for cell culture |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP58050184A JPS59175876A (en) | 1983-03-25 | 1983-03-25 | Medium for cell culture |
Publications (2)
Publication Number | Publication Date |
---|---|
JPS59175876A JPS59175876A (en) | 1984-10-04 |
JPH0543349B2 true JPH0543349B2 (en) | 1993-07-01 |
Family
ID=12852096
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
JP58050184A Granted JPS59175876A (en) | 1983-03-25 | 1983-03-25 | Medium for cell culture |
Country Status (1)
Country | Link |
---|---|
JP (1) | JPS59175876A (en) |
Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JPS59166079A (en) * | 1983-03-14 | 1984-09-19 | Morinaga & Co Ltd | Medium for cell culture |
JPS59173078A (en) * | 1983-03-22 | 1984-09-29 | Morinaga & Co Ltd | Medium for cell culture |
-
1983
- 1983-03-25 JP JP58050184A patent/JPS59175876A/en active Granted
Patent Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JPS59166079A (en) * | 1983-03-14 | 1984-09-19 | Morinaga & Co Ltd | Medium for cell culture |
JPS59173078A (en) * | 1983-03-22 | 1984-09-29 | Morinaga & Co Ltd | Medium for cell culture |
Also Published As
Publication number | Publication date |
---|---|
JPS59175876A (en) | 1984-10-04 |
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