AU2007274065A1 - Cell growth medium - Google Patents
Cell growth medium Download PDFInfo
- Publication number
- AU2007274065A1 AU2007274065A1 AU2007274065A AU2007274065A AU2007274065A1 AU 2007274065 A1 AU2007274065 A1 AU 2007274065A1 AU 2007274065 A AU2007274065 A AU 2007274065A AU 2007274065 A AU2007274065 A AU 2007274065A AU 2007274065 A1 AU2007274065 A1 AU 2007274065A1
- Authority
- AU
- Australia
- Prior art keywords
- cell culture
- embryonic stem
- stem cells
- cell
- primate embryonic
- 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.)
- Abandoned
Links
- 239000001963 growth medium Substances 0.000 title description 4
- 230000010261 cell growth Effects 0.000 title description 2
- 210000004027 cell Anatomy 0.000 claims description 97
- 210000001671 embryonic stem cell Anatomy 0.000 claims description 76
- 238000000034 method Methods 0.000 claims description 72
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 claims description 66
- 238000004113 cell culture Methods 0.000 claims description 58
- 241000288906 Primates Species 0.000 claims description 56
- 239000006143 cell culture medium Substances 0.000 claims description 37
- 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 claims description 36
- HZAXFHJVJLSVMW-UHFFFAOYSA-N 2-Aminoethan-1-ol Chemical compound NCCO HZAXFHJVJLSVMW-UHFFFAOYSA-N 0.000 claims description 35
- 229960005070 ascorbic acid Drugs 0.000 claims description 33
- 235000010323 ascorbic acid Nutrition 0.000 claims description 33
- 239000011668 ascorbic acid Substances 0.000 claims description 33
- 102000018233 Fibroblast Growth Factor Human genes 0.000 claims description 28
- 108050007372 Fibroblast Growth Factor Proteins 0.000 claims description 28
- 229940126864 fibroblast growth factor Drugs 0.000 claims description 28
- 229960002897 heparin Drugs 0.000 claims description 28
- 229920000669 heparin Polymers 0.000 claims description 28
- DGVVWUTYPXICAM-UHFFFAOYSA-N β‐Mercaptoethanol Chemical compound OCCS DGVVWUTYPXICAM-UHFFFAOYSA-N 0.000 claims description 28
- HTTJABKRGRZYRN-UHFFFAOYSA-N Heparin Chemical compound OC1C(NC(=O)C)C(O)OC(COS(O)(=O)=O)C1OC1C(OS(O)(=O)=O)C(O)C(OC2C(C(OS(O)(=O)=O)C(OC3C(C(O)C(O)C(O3)C(O)=O)OS(O)(=O)=O)C(CO)O2)NS(O)(=O)=O)C(C(O)=O)O1 HTTJABKRGRZYRN-UHFFFAOYSA-N 0.000 claims description 27
- WRIDQFICGBMAFQ-UHFFFAOYSA-N (E)-8-Octadecenoic acid Natural products CCCCCCCCCC=CCCCCCCC(O)=O WRIDQFICGBMAFQ-UHFFFAOYSA-N 0.000 claims description 20
- LQJBNNIYVWPHFW-UHFFFAOYSA-N 20:1omega9c fatty acid Natural products CCCCCCCCCCC=CCCCCCCCC(O)=O LQJBNNIYVWPHFW-UHFFFAOYSA-N 0.000 claims description 20
- QSBYPNXLFMSGKH-UHFFFAOYSA-N 9-Heptadecensaeure Natural products CCCCCCCC=CCCCCCCCC(O)=O QSBYPNXLFMSGKH-UHFFFAOYSA-N 0.000 claims description 20
- ZQPPMHVWECSIRJ-UHFFFAOYSA-N Oleic acid Natural products CCCCCCCCC=CCCCCCCCC(O)=O ZQPPMHVWECSIRJ-UHFFFAOYSA-N 0.000 claims description 20
- 239000005642 Oleic acid Substances 0.000 claims description 20
- QXJSBBXBKPUZAA-UHFFFAOYSA-N isooleic acid Natural products CCCCCCCC=CCCCCCCCCC(O)=O QXJSBBXBKPUZAA-UHFFFAOYSA-N 0.000 claims description 20
- ZQPPMHVWECSIRJ-KTKRTIGZSA-N oleic acid Chemical compound CCCCCCCC\C=C/CCCCCCCC(O)=O ZQPPMHVWECSIRJ-KTKRTIGZSA-N 0.000 claims description 20
- 229910019142 PO4 Inorganic materials 0.000 claims description 19
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 claims description 19
- 239000010452 phosphate Substances 0.000 claims description 19
- 108091003079 Bovine Serum Albumin Proteins 0.000 claims description 18
- 108090000379 Fibroblast growth factor 2 Proteins 0.000 claims description 18
- 102000004877 Insulin Human genes 0.000 claims description 18
- 108090001061 Insulin Proteins 0.000 claims description 18
- 229940125396 insulin Drugs 0.000 claims description 18
- 102100024785 Fibroblast growth factor 2 Human genes 0.000 claims description 17
- 102000004338 Transferrin Human genes 0.000 claims description 17
- 108090000901 Transferrin Proteins 0.000 claims description 17
- BVTBRVFYZUCAKH-UHFFFAOYSA-L disodium selenite Chemical compound [Na+].[Na+].[O-][Se]([O-])=O BVTBRVFYZUCAKH-UHFFFAOYSA-L 0.000 claims description 17
- 210000002966 serum Anatomy 0.000 claims description 17
- 239000011781 sodium selenite Substances 0.000 claims description 17
- 235000015921 sodium selenite Nutrition 0.000 claims description 17
- 229960001471 sodium selenite Drugs 0.000 claims description 17
- 239000012581 transferrin Substances 0.000 claims description 17
- 108010085895 Laminin Proteins 0.000 claims description 16
- 102000007547 Laminin Human genes 0.000 claims description 16
- QYSGYZVSCZSLHT-UHFFFAOYSA-N octafluoropropane Chemical compound FC(F)(F)C(F)(F)C(F)(F)F QYSGYZVSCZSLHT-UHFFFAOYSA-N 0.000 claims description 16
- 235000014113 dietary fatty acids Nutrition 0.000 claims description 15
- 239000000194 fatty acid Substances 0.000 claims description 15
- 229930195729 fatty acid Natural products 0.000 claims description 15
- 150000004665 fatty acids Chemical class 0.000 claims description 15
- 102000012422 Collagen Type I Human genes 0.000 claims description 12
- 108010022452 Collagen Type I Proteins 0.000 claims description 12
- 108010035532 Collagen Proteins 0.000 claims description 10
- 102000008186 Collagen Human genes 0.000 claims description 10
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 claims description 10
- 229920001436 collagen Polymers 0.000 claims description 9
- 102100037362 Fibronectin Human genes 0.000 claims description 7
- 108010067306 Fibronectins Proteins 0.000 claims description 7
- 102100035140 Vitronectin Human genes 0.000 claims description 7
- 108010031318 Vitronectin Proteins 0.000 claims description 7
- 230000004069 differentiation Effects 0.000 claims description 7
- 239000003795 chemical substances by application Substances 0.000 claims description 6
- JKMHFZQWWAIEOD-UHFFFAOYSA-N 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid Chemical compound OCC[NH+]1CCN(CCS([O-])(=O)=O)CC1 JKMHFZQWWAIEOD-UHFFFAOYSA-N 0.000 claims description 5
- 239000007995 HEPES buffer Substances 0.000 claims description 5
- 102000029816 Collagenase Human genes 0.000 claims description 4
- 108060005980 Collagenase Proteins 0.000 claims description 4
- 102000004142 Trypsin Human genes 0.000 claims description 4
- 108090000631 Trypsin Proteins 0.000 claims description 4
- 229960002424 collagenase Drugs 0.000 claims description 4
- 239000012588 trypsin Substances 0.000 claims description 4
- 108090000144 Human Proteins Proteins 0.000 claims description 3
- 102000003839 Human Proteins Human genes 0.000 claims description 3
- 102000007056 Recombinant Fusion Proteins Human genes 0.000 claims description 3
- 108010008281 Recombinant Fusion Proteins Proteins 0.000 claims description 3
- 108090000386 Fibroblast Growth Factor 1 Proteins 0.000 claims description 2
- 108050002074 Fibroblast growth factor 17 Proteins 0.000 claims description 2
- 102100035308 Fibroblast growth factor 17 Human genes 0.000 claims description 2
- 108090000381 Fibroblast growth factor 4 Proteins 0.000 claims description 2
- 102100028072 Fibroblast growth factor 4 Human genes 0.000 claims description 2
- 108090000367 Fibroblast growth factor 9 Proteins 0.000 claims description 2
- 102100037665 Fibroblast growth factor 9 Human genes 0.000 claims description 2
- OHJKXVLJWUPWQG-PNRHKHKDSA-N Heparinsodiumsalt Chemical compound O[C@@H]1[C@@H](NS(O)(=O)=O)[C@@H](O)O[C@H](COS(O)(=O)=O)[C@H]1O[C@H]1[C@H](OS(O)(=O)=O)[C@@H](O)[C@H](O)[C@H](C(O)=O)O1 OHJKXVLJWUPWQG-PNRHKHKDSA-N 0.000 claims description 2
- 239000006172 buffering agent Substances 0.000 claims description 2
- 108010007093 dispase Proteins 0.000 claims description 2
- 210000003981 ectoderm Anatomy 0.000 claims description 2
- 210000001900 endoderm Anatomy 0.000 claims description 2
- 210000002919 epithelial cell Anatomy 0.000 claims description 2
- DNJIEGIFACGWOD-UHFFFAOYSA-N ethyl mercaptane Natural products CCS DNJIEGIFACGWOD-UHFFFAOYSA-N 0.000 claims description 2
- 102000003977 fibroblast growth factor 18 Human genes 0.000 claims description 2
- 108090000370 fibroblast growth factor 18 Proteins 0.000 claims description 2
- 210000003716 mesoderm Anatomy 0.000 claims description 2
- 102100031706 Fibroblast growth factor 1 Human genes 0.000 claims 1
- 239000002609 medium Substances 0.000 description 23
- 230000012010 growth Effects 0.000 description 12
- 210000000130 stem cell Anatomy 0.000 description 11
- 210000001519 tissue Anatomy 0.000 description 11
- 210000002950 fibroblast Anatomy 0.000 description 7
- 241001529936 Murinae Species 0.000 description 5
- 239000000243 solution Substances 0.000 description 5
- 241000699666 Mus <mouse, genus> Species 0.000 description 4
- 210000004602 germ cell Anatomy 0.000 description 4
- 102100024505 Bone morphogenetic protein 4 Human genes 0.000 description 3
- 201000009051 Embryonal Carcinoma Diseases 0.000 description 3
- 101000762379 Homo sapiens Bone morphogenetic protein 4 Proteins 0.000 description 3
- 239000002253 acid Substances 0.000 description 3
- 239000007640 basal medium Substances 0.000 description 3
- 230000004663 cell proliferation Effects 0.000 description 3
- 239000003153 chemical reaction reagent Substances 0.000 description 3
- 239000000306 component Substances 0.000 description 3
- 238000011161 development Methods 0.000 description 3
- 230000018109 developmental process Effects 0.000 description 3
- 238000012423 maintenance Methods 0.000 description 3
- 210000004962 mammalian cell Anatomy 0.000 description 3
- 239000004033 plastic Substances 0.000 description 3
- 229920003023 plastic Polymers 0.000 description 3
- 230000000392 somatic effect Effects 0.000 description 3
- JZRWCGZRTZMZEH-UHFFFAOYSA-N thiamine Chemical compound CC1=C(CCO)SC=[N+]1CC1=CN=C(C)N=C1N JZRWCGZRTZMZEH-UHFFFAOYSA-N 0.000 description 3
- YBJHBAHKTGYVGT-ZKWXMUAHSA-N (+)-Biotin Chemical compound N1C(=O)N[C@@H]2[C@H](CCCCC(=O)O)SC[C@@H]21 YBJHBAHKTGYVGT-ZKWXMUAHSA-N 0.000 description 2
- MTCFGRXMJLQNBG-REOHCLBHSA-N (2S)-2-Amino-3-hydroxypropansäure Chemical compound OC[C@H](N)C(O)=O MTCFGRXMJLQNBG-REOHCLBHSA-N 0.000 description 2
- ALYNCZNDIQEVRV-UHFFFAOYSA-N 4-aminobenzoic acid Chemical compound NC1=CC=C(C(O)=O)C=C1 ALYNCZNDIQEVRV-UHFFFAOYSA-N 0.000 description 2
- 241000283690 Bos taurus Species 0.000 description 2
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 2
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 2
- 150000000996 L-ascorbic acids Chemical class 0.000 description 2
- DCXYFEDJOCDNAF-REOHCLBHSA-N L-asparagine Chemical compound OC(=O)[C@@H](N)CC(N)=O DCXYFEDJOCDNAF-REOHCLBHSA-N 0.000 description 2
- HNDVDQJCIGZPNO-YFKPBYRVSA-N L-histidine Chemical compound OC(=O)[C@@H](N)CC1=CN=CN1 HNDVDQJCIGZPNO-YFKPBYRVSA-N 0.000 description 2
- ROHFNLRQFUQHCH-YFKPBYRVSA-N L-leucine Chemical compound CC(C)C[C@H](N)C(O)=O ROHFNLRQFUQHCH-YFKPBYRVSA-N 0.000 description 2
- KDXKERNSBIXSRK-YFKPBYRVSA-N L-lysine Chemical compound NCCCC[C@H](N)C(O)=O KDXKERNSBIXSRK-YFKPBYRVSA-N 0.000 description 2
- COLNVLDHVKWLRT-QMMMGPOBSA-N L-phenylalanine Chemical compound OC(=O)[C@@H](N)CC1=CC=CC=C1 COLNVLDHVKWLRT-QMMMGPOBSA-N 0.000 description 2
- AYFVYJQAPQTCCC-GBXIJSLDSA-N L-threonine Chemical compound C[C@@H](O)[C@H](N)C(O)=O AYFVYJQAPQTCCC-GBXIJSLDSA-N 0.000 description 2
- QIVBCDIJIAJPQS-VIFPVBQESA-N L-tryptophane Chemical compound C1=CC=C2C(C[C@H](N)C(O)=O)=CNC2=C1 QIVBCDIJIAJPQS-VIFPVBQESA-N 0.000 description 2
- OUYCCCASQSFEME-QMMMGPOBSA-N L-tyrosine Chemical compound OC(=O)[C@@H](N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-QMMMGPOBSA-N 0.000 description 2
- KZSNJWFQEVHDMF-BYPYZUCNSA-N L-valine Chemical compound CC(C)[C@H](N)C(O)=O KZSNJWFQEVHDMF-BYPYZUCNSA-N 0.000 description 2
- TWRXJAOTZQYOKJ-UHFFFAOYSA-L Magnesium chloride Chemical compound [Mg+2].[Cl-].[Cl-] TWRXJAOTZQYOKJ-UHFFFAOYSA-L 0.000 description 2
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 2
- WCUXLLCKKVVCTQ-UHFFFAOYSA-M Potassium chloride Chemical compound [Cl-].[K+] WCUXLLCKKVVCTQ-UHFFFAOYSA-M 0.000 description 2
- 102000029797 Prion Human genes 0.000 description 2
- 108091000054 Prion Proteins 0.000 description 2
- RADKZDMFGJYCBB-UHFFFAOYSA-N Pyridoxal Chemical compound CC1=NC=C(CO)C(C=O)=C1O RADKZDMFGJYCBB-UHFFFAOYSA-N 0.000 description 2
- AUNGANRZJHBGPY-SCRDCRAPSA-N Riboflavin Chemical compound OC[C@@H](O)[C@@H](O)[C@@H](O)CN1C=2C=C(C)C(C)=CC=2N=C2C1=NC(=O)NC2=O AUNGANRZJHBGPY-SCRDCRAPSA-N 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- QAOWNCQODCNURD-UHFFFAOYSA-L Sulfate Chemical compound [O-]S([O-])(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-L 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
- 241000700605 Viruses Species 0.000 description 2
- ZCCIPPOKBCJFDN-UHFFFAOYSA-N calcium nitrate Chemical compound [Ca+2].[O-][N+]([O-])=O.[O-][N+]([O-])=O ZCCIPPOKBCJFDN-UHFFFAOYSA-N 0.000 description 2
- 229910002091 carbon monoxide Inorganic materials 0.000 description 2
- 238000012258 culturing Methods 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 210000002257 embryonic structure Anatomy 0.000 description 2
- 239000012091 fetal bovine serum Substances 0.000 description 2
- OVBPIULPVIDEAO-LBPRGKRZSA-N folic acid Chemical compound C=1N=C2NC(N)=NC(=O)C2=NC=1CNC1=CC=C(C(=O)N[C@@H](CCC(O)=O)C(O)=O)C=C1 OVBPIULPVIDEAO-LBPRGKRZSA-N 0.000 description 2
- BTCSSZJGUNDROE-UHFFFAOYSA-N gamma-aminobutyric acid Chemical compound NCCCC(O)=O BTCSSZJGUNDROE-UHFFFAOYSA-N 0.000 description 2
- 239000008103 glucose Substances 0.000 description 2
- RWSXRVCMGQZWBV-WDSKDSINSA-N glutathione Chemical compound OC(=O)[C@@H](N)CCC(=O)N[C@@H](CS)C(=O)NCC(O)=O RWSXRVCMGQZWBV-WDSKDSINSA-N 0.000 description 2
- -1 heparin sulphate salt Chemical class 0.000 description 2
- FDGQSTZJBFJUBT-UHFFFAOYSA-N hypoxanthine Chemical compound O=C1NC=NC2=C1NC=N2 FDGQSTZJBFJUBT-UHFFFAOYSA-N 0.000 description 2
- 238000000338 in vitro Methods 0.000 description 2
- VCJMYUPGQJHHFU-UHFFFAOYSA-N iron(3+);trinitrate Chemical compound [Fe+3].[O-][N+]([O-])=O.[O-][N+]([O-])=O.[O-][N+]([O-])=O VCJMYUPGQJHHFU-UHFFFAOYSA-N 0.000 description 2
- 210000001161 mammalian embryo Anatomy 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 210000002894 multi-fate stem cell Anatomy 0.000 description 2
- 210000001778 pluripotent stem cell Anatomy 0.000 description 2
- LXNHXLLTXMVWPM-UHFFFAOYSA-N pyridoxine Chemical compound CC1=NC=C(CO)C(CO)=C1O LXNHXLLTXMVWPM-UHFFFAOYSA-N 0.000 description 2
- 230000008672 reprogramming Effects 0.000 description 2
- DAEPDZWVDSPTHF-UHFFFAOYSA-M sodium pyruvate Chemical compound [Na+].CC(=O)C([O-])=O DAEPDZWVDSPTHF-UHFFFAOYSA-M 0.000 description 2
- 229910021653 sulphate ion Inorganic materials 0.000 description 2
- 239000000725 suspension Substances 0.000 description 2
- OGNSCSPNOLGXSM-UHFFFAOYSA-N (+/-)-DABA Natural products NCCC(N)C(O)=O OGNSCSPNOLGXSM-UHFFFAOYSA-N 0.000 description 1
- HHGZUQPEIHGQST-RGVONZFCSA-N (2r)-2-amino-3-[[(2r)-2-amino-2-carboxyethyl]disulfanyl]propanoic acid;dihydrochloride Chemical compound Cl.Cl.OC(=O)[C@@H](N)CSSC[C@H](N)C(O)=O HHGZUQPEIHGQST-RGVONZFCSA-N 0.000 description 1
- 239000001763 2-hydroxyethyl(trimethyl)azanium Substances 0.000 description 1
- 108010088751 Albumins Proteins 0.000 description 1
- 102000009027 Albumins Human genes 0.000 description 1
- 102000002260 Alkaline Phosphatase Human genes 0.000 description 1
- 108020004774 Alkaline Phosphatase Proteins 0.000 description 1
- KWTQSFXGGICVPE-WCCKRBBISA-N Arginine hydrochloride Chemical compound Cl.OC(=O)[C@@H](N)CCCN=C(N)N KWTQSFXGGICVPE-WCCKRBBISA-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
- 241000537222 Betabaculovirus Species 0.000 description 1
- UXVMQQNJUSDDNG-UHFFFAOYSA-L Calcium chloride Chemical compound [Cl-].[Cl-].[Ca+2] UXVMQQNJUSDDNG-UHFFFAOYSA-L 0.000 description 1
- 235000019743 Choline chloride Nutrition 0.000 description 1
- AUNGANRZJHBGPY-UHFFFAOYSA-N D-Lyxoflavin Natural products OCC(O)C(O)C(O)CN1C=2C=C(C)C(C)=CC=2N=C2C1=NC(=O)NC2=O AUNGANRZJHBGPY-UHFFFAOYSA-N 0.000 description 1
- QNAYBMKLOCPYGJ-UHFFFAOYSA-N D-alpha-Ala Natural products CC([NH3+])C([O-])=O QNAYBMKLOCPYGJ-UHFFFAOYSA-N 0.000 description 1
- QXNVGIXVLWOKEQ-UHFFFAOYSA-N Disodium Chemical compound [Na][Na] QXNVGIXVLWOKEQ-UHFFFAOYSA-N 0.000 description 1
- 239000012591 Dulbecco’s Phosphate Buffered Saline Substances 0.000 description 1
- YCAGGFXSFQFVQL-UHFFFAOYSA-N Endothion Chemical compound COC1=COC(CSP(=O)(OC)OC)=CC1=O YCAGGFXSFQFVQL-UHFFFAOYSA-N 0.000 description 1
- 102000003971 Fibroblast Growth Factor 1 Human genes 0.000 description 1
- 102000003974 Fibroblast growth factor 2 Human genes 0.000 description 1
- 239000001828 Gelatine Substances 0.000 description 1
- 108010024636 Glutathione Proteins 0.000 description 1
- 239000004471 Glycine Substances 0.000 description 1
- SQUHHTBVTRBESD-UHFFFAOYSA-N Hexa-Ac-myo-Inositol Natural products CC(=O)OC1C(OC(C)=O)C(OC(C)=O)C(OC(C)=O)C(OC(C)=O)C1OC(C)=O SQUHHTBVTRBESD-UHFFFAOYSA-N 0.000 description 1
- PMMYEEVYMWASQN-DMTCNVIQSA-N Hydroxyproline Chemical compound O[C@H]1CN[C@H](C(O)=O)C1 PMMYEEVYMWASQN-DMTCNVIQSA-N 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
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 description 1
- QNAYBMKLOCPYGJ-UWTATZPHSA-N L-Alanine Natural products C[C@@H](N)C(O)=O QNAYBMKLOCPYGJ-UWTATZPHSA-N 0.000 description 1
- 235000019766 L-Lysine Nutrition 0.000 description 1
- FFEARJCKVFRZRR-UHFFFAOYSA-N L-Methionine Natural products CSCCC(N)C(O)=O FFEARJCKVFRZRR-UHFFFAOYSA-N 0.000 description 1
- ONIBWKKTOPOVIA-BYPYZUCNSA-N L-Proline Chemical compound OC(=O)[C@@H]1CCCN1 ONIBWKKTOPOVIA-BYPYZUCNSA-N 0.000 description 1
- QNAYBMKLOCPYGJ-REOHCLBHSA-N L-alanine Chemical compound C[C@H](N)C(O)=O QNAYBMKLOCPYGJ-REOHCLBHSA-N 0.000 description 1
- ODKSFYDXXFIFQN-BYPYZUCNSA-N L-arginine Chemical compound OC(=O)[C@@H](N)CCCN=C(N)N ODKSFYDXXFIFQN-BYPYZUCNSA-N 0.000 description 1
- 229930064664 L-arginine Natural products 0.000 description 1
- 235000014852 L-arginine Nutrition 0.000 description 1
- IFQSXNOEEPCSLW-DKWTVANSSA-N L-cysteine hydrochloride Chemical compound Cl.SC[C@H](N)C(O)=O IFQSXNOEEPCSLW-DKWTVANSSA-N 0.000 description 1
- ZDXPYRJPNDTMRX-VKHMYHEASA-N L-glutamine Chemical compound OC(=O)[C@@H](N)CCC(N)=O ZDXPYRJPNDTMRX-VKHMYHEASA-N 0.000 description 1
- 229930182816 L-glutamine Natural products 0.000 description 1
- AGPKZVBTJJNPAG-WHFBIAKZSA-N L-isoleucine Chemical compound CC[C@H](C)[C@H](N)C(O)=O AGPKZVBTJJNPAG-WHFBIAKZSA-N 0.000 description 1
- 229930182844 L-isoleucine Natural products 0.000 description 1
- 239000004395 L-leucine Substances 0.000 description 1
- 235000019454 L-leucine Nutrition 0.000 description 1
- FFEARJCKVFRZRR-BYPYZUCNSA-N L-methionine Chemical compound CSCC[C@H](N)C(O)=O FFEARJCKVFRZRR-BYPYZUCNSA-N 0.000 description 1
- 229930195722 L-methionine Natural products 0.000 description 1
- 229930182821 L-proline Natural products 0.000 description 1
- OYHQOLUKZRVURQ-HZJYTTRNSA-N Linoleic acid Chemical compound CCCCC\C=C/C\C=C/CCCCCCCC(O)=O OYHQOLUKZRVURQ-HZJYTTRNSA-N 0.000 description 1
- WHXSMMKQMYFTQS-UHFFFAOYSA-N Lithium Chemical compound [Li] WHXSMMKQMYFTQS-UHFFFAOYSA-N 0.000 description 1
- 239000004472 Lysine Substances 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 229930192392 Mitomycin Natural products 0.000 description 1
- 241000699670 Mus sp. Species 0.000 description 1
- NWIBSHFKIJFRCO-WUDYKRTCSA-N Mytomycin Chemical compound C1N2C(C(C(C)=C(N)C3=O)=O)=C3[C@@H](COC(N)=O)[C@@]2(OC)[C@@H]2[C@H]1N2 NWIBSHFKIJFRCO-WUDYKRTCSA-N 0.000 description 1
- OVBPIULPVIDEAO-UHFFFAOYSA-N N-Pteroyl-L-glutaminsaeure Natural products C=1N=C2NC(N)=NC(=O)C2=NC=1CNC1=CC=C(C(=O)NC(CCC(O)=O)C(O)=O)C=C1 OVBPIULPVIDEAO-UHFFFAOYSA-N 0.000 description 1
- DFPAKSUCGFBDDF-UHFFFAOYSA-N Nicotinamide Chemical compound NC(=O)C1=CC=CN=C1 DFPAKSUCGFBDDF-UHFFFAOYSA-N 0.000 description 1
- BELBBZDIHDAJOR-UHFFFAOYSA-N Phenolsulfonephthalein Chemical compound C1=CC(O)=CC=C1C1(C=2C=CC(O)=CC=2)C2=CC=CC=C2S(=O)(=O)O1 BELBBZDIHDAJOR-UHFFFAOYSA-N 0.000 description 1
- NBIIXXVUZAFLBC-UHFFFAOYSA-L Phosphate ion(2-) Chemical compound OP([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-L 0.000 description 1
- 206010043276 Teratoma Diseases 0.000 description 1
- 239000004473 Threonine Substances 0.000 description 1
- 230000002378 acidificating effect Effects 0.000 description 1
- 239000000654 additive Substances 0.000 description 1
- 210000004504 adult stem cell Anatomy 0.000 description 1
- 229960003767 alanine Drugs 0.000 description 1
- 229940024606 amino acid Drugs 0.000 description 1
- 235000001014 amino acid Nutrition 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 108010054176 apotransferrin Proteins 0.000 description 1
- 229960001230 asparagine Drugs 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
- 230000015572 biosynthetic process Effects 0.000 description 1
- 229960002685 biotin Drugs 0.000 description 1
- 235000020958 biotin Nutrition 0.000 description 1
- 239000011616 biotin Substances 0.000 description 1
- 210000002459 blastocyst Anatomy 0.000 description 1
- 239000011575 calcium Substances 0.000 description 1
- FAPWYRCQGJNNSJ-UBKPKTQASA-L calcium D-pantothenic acid Chemical compound [Ca+2].OCC(C)(C)[C@@H](O)C(=O)NCCC([O-])=O.OCC(C)(C)[C@@H](O)C(=O)NCCC([O-])=O FAPWYRCQGJNNSJ-UBKPKTQASA-L 0.000 description 1
- 239000001110 calcium chloride Substances 0.000 description 1
- 229910001628 calcium chloride Inorganic materials 0.000 description 1
- 229960002079 calcium pantothenate Drugs 0.000 description 1
- 239000012534 cell culture medium component Substances 0.000 description 1
- 230000007910 cell fusion Effects 0.000 description 1
- 239000006285 cell suspension Substances 0.000 description 1
- SGMZJAMFUVOLNK-UHFFFAOYSA-M choline chloride Chemical compound [Cl-].C[N+](C)(C)CCO SGMZJAMFUVOLNK-UHFFFAOYSA-M 0.000 description 1
- 229960003178 choline chloride Drugs 0.000 description 1
- 150000001875 compounds Chemical group 0.000 description 1
- 229910000365 copper sulfate Inorganic materials 0.000 description 1
- ARUVKPQLZAKDPS-UHFFFAOYSA-L copper(II) sulfate Chemical compound [Cu+2].[O-][S+2]([O-])([O-])[O-] ARUVKPQLZAKDPS-UHFFFAOYSA-L 0.000 description 1
- 238000012136 culture method Methods 0.000 description 1
- 210000000805 cytoplasm Anatomy 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 238000007877 drug screening Methods 0.000 description 1
- 210000002242 embryoid body Anatomy 0.000 description 1
- 210000003527 eukaryotic cell Anatomy 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 239000011790 ferrous sulphate Substances 0.000 description 1
- 235000003891 ferrous sulphate Nutrition 0.000 description 1
- 229960000304 folic acid Drugs 0.000 description 1
- 235000019152 folic acid Nutrition 0.000 description 1
- 239000011724 folic acid Substances 0.000 description 1
- 229960003692 gamma aminobutyric acid Drugs 0.000 description 1
- 229920000159 gelatin Polymers 0.000 description 1
- 235000019322 gelatine Nutrition 0.000 description 1
- 210000004392 genitalia Anatomy 0.000 description 1
- 229960003180 glutathione Drugs 0.000 description 1
- 229960002885 histidine Drugs 0.000 description 1
- 239000012678 infectious agent Substances 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 229960000367 inositol Drugs 0.000 description 1
- CDAISMWEOUEBRE-GPIVLXJGSA-N inositol Chemical compound O[C@H]1[C@H](O)[C@@H](O)[C@H](O)[C@H](O)[C@@H]1O CDAISMWEOUEBRE-GPIVLXJGSA-N 0.000 description 1
- BAUYGSIQEAFULO-UHFFFAOYSA-L iron(2+) sulfate (anhydrous) Chemical compound [Fe+2].[O-]S([O-])(=O)=O BAUYGSIQEAFULO-UHFFFAOYSA-L 0.000 description 1
- 229910000359 iron(II) sulfate Inorganic materials 0.000 description 1
- 229960000310 isoleucine Drugs 0.000 description 1
- 239000010410 layer Substances 0.000 description 1
- 229960003136 leucine Drugs 0.000 description 1
- 208000032839 leukemia Diseases 0.000 description 1
- 235000020778 linoleic acid Nutrition 0.000 description 1
- OYHQOLUKZRVURQ-IXWMQOLASA-N linoleic acid Natural products CCCCC\C=C/C\C=C\CCCCCCCC(O)=O OYHQOLUKZRVURQ-IXWMQOLASA-N 0.000 description 1
- 150000002632 lipids Chemical class 0.000 description 1
- AGBQKNBQESQNJD-UHFFFAOYSA-M lipoate Chemical compound [O-]C(=O)CCCCC1CCSS1 AGBQKNBQESQNJD-UHFFFAOYSA-M 0.000 description 1
- 235000019136 lipoic acid Nutrition 0.000 description 1
- 229910052744 lithium Inorganic materials 0.000 description 1
- 239000011777 magnesium Substances 0.000 description 1
- 229910001629 magnesium chloride Inorganic materials 0.000 description 1
- 229910052943 magnesium sulfate Inorganic materials 0.000 description 1
- 235000019341 magnesium sulphate Nutrition 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 108010082117 matrigel Proteins 0.000 description 1
- 210000000713 mesentery Anatomy 0.000 description 1
- 229960004452 methionine Drugs 0.000 description 1
- 230000002297 mitogenic effect Effects 0.000 description 1
- 229960004857 mitomycin Drugs 0.000 description 1
- XONPDZSGENTBNJ-UHFFFAOYSA-N molecular hydrogen;sodium Chemical compound [Na].[H][H] XONPDZSGENTBNJ-UHFFFAOYSA-N 0.000 description 1
- 238000004264 monolayer culture Methods 0.000 description 1
- 210000002569 neuron Anatomy 0.000 description 1
- 230000004031 neuronal differentiation Effects 0.000 description 1
- 239000002547 new drug Substances 0.000 description 1
- 229960003966 nicotinamide Drugs 0.000 description 1
- 235000005152 nicotinamide Nutrition 0.000 description 1
- 239000011570 nicotinamide Substances 0.000 description 1
- 210000000287 oocyte Anatomy 0.000 description 1
- 230000003647 oxidation Effects 0.000 description 1
- 238000007254 oxidation reaction Methods 0.000 description 1
- 230000002974 pharmacogenomic effect Effects 0.000 description 1
- 229960003531 phenolsulfonphthalein Drugs 0.000 description 1
- 229960005190 phenylalanine Drugs 0.000 description 1
- 210000002826 placenta Anatomy 0.000 description 1
- 239000001103 potassium chloride Substances 0.000 description 1
- 235000011164 potassium chloride Nutrition 0.000 description 1
- 239000002243 precursor Substances 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- 229960002429 proline Drugs 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 229960003581 pyridoxal Drugs 0.000 description 1
- 235000008164 pyridoxal Nutrition 0.000 description 1
- 239000011674 pyridoxal Substances 0.000 description 1
- 235000008160 pyridoxine Nutrition 0.000 description 1
- 239000011677 pyridoxine Substances 0.000 description 1
- 229960002477 riboflavin Drugs 0.000 description 1
- 235000019192 riboflavin Nutrition 0.000 description 1
- 239000002151 riboflavin Substances 0.000 description 1
- CDAISMWEOUEBRE-UHFFFAOYSA-N scyllo-inosotol Natural products OC1C(O)C(O)C(O)C(O)C1O CDAISMWEOUEBRE-UHFFFAOYSA-N 0.000 description 1
- 229940082569 selenite Drugs 0.000 description 1
- MCAHWIHFGHIESP-UHFFFAOYSA-L selenite(2-) Chemical compound [O-][Se]([O-])=O MCAHWIHFGHIESP-UHFFFAOYSA-L 0.000 description 1
- 229960001153 serine Drugs 0.000 description 1
- 239000011734 sodium Substances 0.000 description 1
- 229910052708 sodium Inorganic materials 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- 229940054269 sodium pyruvate Drugs 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 239000000126 substance Chemical group 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 230000000153 supplemental effect Effects 0.000 description 1
- 208000001608 teratocarcinoma Diseases 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
- 238000002560 therapeutic procedure Methods 0.000 description 1
- 235000019157 thiamine Nutrition 0.000 description 1
- 229960003495 thiamine Drugs 0.000 description 1
- 239000011721 thiamine Substances 0.000 description 1
- 229960002663 thioctic acid Drugs 0.000 description 1
- 229960002898 threonine Drugs 0.000 description 1
- 229940104230 thymidine Drugs 0.000 description 1
- 210000003014 totipotent stem cell Anatomy 0.000 description 1
- 231100000027 toxicology Toxicity 0.000 description 1
- FGMPLJWBKKVCDB-UHFFFAOYSA-N trans-L-hydroxy-proline Natural products ON1CCCC1C(O)=O FGMPLJWBKKVCDB-UHFFFAOYSA-N 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 229960004799 tryptophan Drugs 0.000 description 1
- 229960004441 tyrosine Drugs 0.000 description 1
- 229960004295 valine Drugs 0.000 description 1
- 229940088594 vitamin Drugs 0.000 description 1
- 229930003231 vitamin Natural products 0.000 description 1
- 235000013343 vitamin Nutrition 0.000 description 1
- 239000011782 vitamin Substances 0.000 description 1
- 229940011671 vitamin b6 Drugs 0.000 description 1
- 239000002676 xenobiotic agent Substances 0.000 description 1
- 230000002034 xenobiotic effect Effects 0.000 description 1
- NWONKYPBYAMBJT-UHFFFAOYSA-L zinc sulfate Chemical compound [Zn+2].[O-]S([O-])(=O)=O NWONKYPBYAMBJT-UHFFFAOYSA-L 0.000 description 1
- 229910000368 zinc sulfate Inorganic materials 0.000 description 1
- 229960001763 zinc sulfate Drugs 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
- C12N5/0603—Embryonic cells ; Embryoid bodies
- C12N5/0606—Pluripotent embryonic cells, e.g. embryonic stem cells [ES]
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2500/00—Specific components of cell culture medium
- C12N2500/05—Inorganic components
- C12N2500/10—Metals; Metal chelators
- C12N2500/20—Transition metals
- C12N2500/24—Iron; Fe chelators; Transferrin
- C12N2500/25—Insulin-transferrin; Insulin-transferrin-selenium
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2500/00—Specific components of cell culture medium
- C12N2500/30—Organic components
- C12N2500/38—Vitamins
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2500/00—Specific components of cell culture medium
- C12N2500/30—Organic components
- C12N2500/44—Thiols, e.g. mercaptoethanol
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2500/00—Specific components of cell culture medium
- C12N2500/30—Organic components
- C12N2500/46—Amines, e.g. putrescine
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2500/00—Specific components of cell culture medium
- C12N2500/90—Serum-free medium, which may still contain naturally-sourced components
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/10—Growth factors
- C12N2501/115—Basic fibroblast growth factor (bFGF, FGF-2)
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2501/00—Active agents used in cell culture processes, e.g. differentation
- C12N2501/90—Polysaccharides
- C12N2501/91—Heparin
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Biomedical Technology (AREA)
- Genetics & Genomics (AREA)
- Zoology (AREA)
- Gynecology & Obstetrics (AREA)
- Biotechnology (AREA)
- Organic Chemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Developmental Biology & Embryology (AREA)
- Chemical & Material Sciences (AREA)
- Wood Science & Technology (AREA)
- Reproductive Health (AREA)
- Microbiology (AREA)
- Biochemistry (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Cell Biology (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Apparatus Associated With Microorganisms And Enzymes (AREA)
Description
WO 2008/007082 PCT/GB2007/002584 Cell Growth Medium The invention relates to the maintenance of primate embryonic stem cells, preferably human embryonic stem cells (hES), in culture in the absence of feeder cells and serum. 5 The culturing of eukaryotic cells, for example some mammalian cells has become a routine procedure and cell culture conditions which allow certain cells to proliferate are well defined. Typically, cell culture of mammalian cells requires a sterile vessel, usually manufactured from plastics and growth medium. The growth of, for example embryonic 10 stem cells requires the presence of both feeder cells and serum. The function of the feeder cells is not known with certainty. However, it is speculated that feeder cells may function to provide mitogenic signals which stimulate cell proliferation and/or maintain cells in an undifferentiated state. Feeder cells are typically fibroblasts which have been treated such that they cannot proliferate (e.g. mitomycin or irradiation treatment). Typically, feeder 15 fibroblasts are murine in origin but may be derived from other species The term "stem cell" represents a generic group of undifferentiated cells that possess the capacity for self-renewal while retaining varying potentials to form differentiated cells and tissues. Stem cells can be totipotent, pluripotent or multipotent. Derivative stem cells that 20 have lost the ability to differentiate also occur and are termed 'nullipotent' stem cells. A totipotent stem cell is a cell that has the ability to form all the cells and tissues that are found in an intact organism, including the extra-embryonic tissues (i.e. the placenta). Totipotent cells comprise the very early embryo (8 cells) and have the ability to form an intact organism. A pluripotent stem cell is a cell that has the ability to form all tissues 25 found in an intact organism although the pluripotent stem cell cannot form an intact organism. A multipotent cell has a restricted ability to form differentiated cells and tissues. Typically adult stem cells are multipotent stem cells and are the precursor stem cells or lineage restricted stem cells that have the ability to form some cells or tissues and replenish senescing or damaged cells/tissues. Generally they cannot form all tissues 30 found in an organism, although some reports have claimed a greater potential for such 'adult' stem cells than originally thought. Pluripotent embryonic stem cells may be principally derived from two embryonic sources. 1 WO 2008/007082 PCT/GB2007/002584 Cells isolated from the inner cell mass are termed embryonic stem (ES) cells. In the laboratory mouse, similar cells can be derived from the culture of primordial germ cells isolated from the mesenteries or genital ridges of days 8.5-12.5 post coitum embryos. These are referred to as embryonic germ cells (EG cells). Each of these types of 5 pluripotential cell has a similar developmental potential with respect to differentiation into alternate cell types, but possible differences in behaviour (e.g. with respect to imprinting) have led these cells to be distinguished from one another. However, the term "pluripotent embryonic stem cell" encompasses both cells derived from the inner cell mass and primordial germ cells. 10 The establishment of in vitro cultures of primate embryonic stem cells has proven to be problematic. An indication that conditions may be determined which could allow the establishment of hES cells in culture is described in W096/22362. W096/22362 describes cell lines and growth conditions which allow the continuous proliferation of 15 primate ES cells which exhibit a range of characteristics or markers which are associated with stem cells having pluripotent characteristics. These include, but are not limited to; maintenance in culture for at least 20 passages when maintained on fibroblast feeder layers; production of clusters of cells referred to as embryoid bodies; when cultured in suspension, an ability to differentiate into multiple cell types in monolayer culture; the 20 formation of xenograft teratomas with multiple differentiated cell types when injected into immunodeficient mice, and the expression of embryonic stem cell specific markers, notably SSEA3, SSEA4, TRA-1-60, TRA-1-81, alkaline phosphatase, and Oct4. W096/22362 discloses a method of maintaining primate ES cells in culture in an undifferentiated state in the presence of mouse fibroblast feeder cells and serum. 25 The potential utility of embryonic stem cells, particularly human ES (hES) cells, in therapeutic tissue engineering is well documented. The pluripotent nature of these cells enables the selection and differentiation of hES cells into any cell/tissue type. However, the potential risk is that adventitious agents such as prions or viruses may infect the 30 recipient when cells exposed to fetal bovine serum or murine feeder cells are used in therapy. It is therefore essential that cell culture of hES cells is conducted to minimise this risk. The development of feeder free and serum free conditions will help reduce this risk. 2 WO 2008/007082 PCT/GB2007/002584 Moreover, hES cells that have been differentiated into particular cell type derivatives have utility in the identification gene targets for new drugs and existing drugs since the cells are genotypically identical, stable and of known origin. The use of ES cell lines of distinct genotypes also offers possible routes to drug screening and toxicology in a way pertinent 5 to pharmacogenomics. The development of serum free conditions for the culture of primate ES cells is known. For example, WO01/66697 discloses serum free growth of primate ES cells wherein the serum is replaced with fibroblast growth factor, typically human basic fibroblast growth 10 factor (bFGF 4ng/ml). The cell culture media includes KnockOut SR" (described in WO98/30679 which is incorporated by reference in its entirety) supplemented with bFGF. However the cell culture includes irradiated murine fibroblast feeder cells. The development of a serum free and feeder free culture method for the growth of hES 15 cells is disclosed in W02006/029198. These growth conditions use elevated concentrations of bFGF (40-100ng/ml), supplemental agents that include gamma amino butyric acid, pipecholic acid and lithium and including amino acids, lipids, vitamins and glucose. W02006/029198 also discloses the use of a cell culture substrate comprising human proteins such as fibronectin, vitronectin and laminin. 20 Furthermore, Furue et al (In vitro Cell Dev. Biol. Animal 41:19-28, 2005) discloses the serum and feeder free growth of mouse embryonic stem cells in the presence of leukaemia inhibitory factor (LIF). This is also described in WO2005/063968. 25 It would be advantageous if simple cell culture conditions could be established which did not require the addition of xenobiotic materials such as fetal bovine serum or murine feeder cells since their use increases the likelihood of infectious agents (e.g. viruses and prions, in particular for bovine products, and murine viruses for mouse feeder cells) infecting mammalian cells grown in culture. The present disclosure provides an 30 alternative simple cell culture medium that allows the maintenance of hES cells under serum and feeder free conditions. 3 WO 2008/007082 PCT/GB2007/002584 According to an aspect of the invention there is provided a method to maintain a primate embryonic stem cell in cell culture conditions that are cell feeder and serum free comprising: forming a preparation of primate embryonic stem cells in a cell culture vessel comprising cell culture medium that includes fibroblast growth factor, heparin and 5 ascorbic acid, or ascorbic acid phosphate, or derivative thereof and maintaining the primate embryonic stem cells in an undifferentiated state. According to an aspect of the invention there is provide a method to maintain primate embryonic stem cells in cell culture conditions that are cell feeder free and serum free 10 comprising the steps: i) forming a preparation of primate embryonic stem cells in a cell culture vessel which is coated with a proteinaceous based cell culture support wherein said cells are cultured in a cell culture medium comprising: insulin, transferrin, sodium selenite, ethanolamine, 2-mercaptoethanol, oleic acid complexed with fatty acid free bovine 15 albumin and further wherein the cell culture medium is supplemented with fibroblast growth factor, heparin and ascorbic acid, or ascorbic acid phosphate, or derivative thereof; and ii) maintaining the primate embryonic stem cells in an undifferentiated state. 20 This disclosure encompasses primate, in particular human, pluripotent embryonic stem cells and also teratocarcinoma stem cells, known as embryonal carcinoma (EC) cells. "Pluripotent embryonic stem cells" relates to both cells derived from the inner cell mass and primordial germ cells (EG). The possibility also exists of reprogramming somatic or 25 extraembryonic differentiated cells, or more restricted stem cells back to a pluripotent state resembling that of ES cells derived from early embryos. One way in which this may be achieved is by somatic nuclear transfer of a nucleus from such a differentiated cell into an enucleated oocyte which is then stimulated to develop as an embryo to the blastocyst stage from which ES cell lines are then derived. Experiments with cell fusion also 30 indicate that the cytoplasm of EC and ES cells may also be capable of reprogramming somatic and other cell types back to an ES-like state. In a preferred method of the invention said cells have a stable karyotype. 4 WO 2008/007082 PCT/GB2007/002584 In a further preferred method of the invention ascorbic acid is ascorbic acid phosphate. 5 Functional derivatives of ascorbic acid and ascorbic acid phosphate are known in the art. For example, EP1666484 the content of which is incorporated by reference in its entirety describes stable derivatives of ascorbic acid which exhibit increased stability to heat or oxidation. 10 In a preferred method of the invention said primate embryonic stem cells are pluripotent human embryonic stem cells. In a preferred embodiment of the invention said primate embryonic stem cells retain the property to differentiate into at least the endoderm, mesoderm and ectoderm tissues 15 throughout cell culture. In a further preferred method of the invention fibroblast growth factor (FGF) is selected from the group consisting of: bFGF/FGF-2, hereinafter acidic FGF/FGF-1, bFGF, FGF-4, FGF-9, FGF-17 or FGF-18. 20 In a preferred method of the invention said fibroblast growth factor is bFGF. Preferably, bFGF is provided at a concentration of between 1-50ng/ml; preferably about 10 ng/ml. Preferably fibroblast growth factor is recombinant. 25 In a further preferred method of the invention ascorbic acid phosphate is provided at a concentration of 10-300pLg/ml; preferably about 100tg/ml. In a further preferred method of the invention 2-ethanolamine is provided at a 30 concentration of 0.05-2.0gg/ml; preferably about 0.6 jtg/ml. In a further preferred method of the invention oleic acid is provided at a concentration of 3-15p.g/ml; preferably about 9.5pg/ml. 5 WO 2008/007082 PCT/GB2007/002584 In a further preferred method of the invention heparin is provided at a concentration of 10 500ng/ml; preferably about 100ng/ml; preferably, heparin is heparin sulphate salt. 5 In a preferred method of the invention said proteinaceous cell culture support is collagen based. In a preferred method of the invention the collagen-based cell culture support comprises 10 type I collagen; preferably recombinant type I collagen. In an alternative preferred method of the invention said cell culture support comprises recombinant human proteins selected from the group consisting of: collagen I, collagen IV, fibronectin, laminin and vitronectin. 15 In a preferred method of the invention said cell support comprises at least two recombinant proteins selected from the group consisting of: collagen I, collagen IV, fibronectin, laminin and vitronectin. 20 In a preferred method of the invention said cell support comprises the recombinant proteins collagen I, collagen IV, fibronectin, laminin and vitronectin. In a preferred method of the invention said cell culture support is Matrigel'. 25 In a preferred method of the invention said primate embryonic stem cells are passaged after addition of EDTA to the cell culture vessel. In an alternative preferred method of the invention said primate embryonic stem cells are passaged after addition of collagenase, preferably collagenase IV. 30 In an alternative preferred method of the invention said primate embryonic stem cells are passaged after addition of dispase. 6 WO 2008/007082 PCT/GB2007/002584 In an alternative preferred method of the invention said primate embryonic stem cells are passaged after addition of trypsin/EDTA, preferably recombinant trypsin. In a further preferred method of the invention said primate embryonic stem cells are 5 cloned. In a preferred method of the invention the cell culture media does not include the buffering agent HEPES. 10 According to a further aspect of the invention there is provide a method to differentiate primate embryonic stem cells into at least one cell-type in cell culture conditions that are cell feeder free and serum free comprising the steps: i) forming a preparation of primate embryonic stem cells in a cell culture vessel which is coated with a proteinaceous based cell culture support wherein said cells are 15 cultured in a cell culture medium comprising: insulin, transferrin, sodium selenite, ethanolamine, 2-mercaptoethanol, oleic acid complexed with fatty acid free bovine albumin and further wherein the cell culture medium is supplemented with fibroblast growth factor and heparin; and ii) adding an agent that induces the differentiation of the primate embryonic stem 20 cells into at least one cell-type. In a preferred method of the invention the primate embryonic stem cells are human pluripotent embryonic stem cells. 25 In a preferred method of the invention the cell-type is a neurone. In an alternative method of the invention the cell-type is an epithelial cell. In a preferred method of the invention said proteinaceous based cell culture support is 30 laminin. According to a further aspect of the invention there is provided a cell culture comprising: primate embryonic stem cells on a proteinaceous based cell culture support and cell 7 WO 2008/007082 PCT/GB2007/002584 culture media comprising: insulin, transferrin, sodium selenite, ethanolamine, 2 mercaptoethanol, oleic acid complexed with fatty acid free bovine albumin and further wherein the cell culture medium is supplemented with fibroblast growth factor, heparin and ascorbic acid, or ascorbic acid phosphate, or derivative thereof. 5 In a preferred embodiment of the invention the primate embryonic stem cells are pluripotent human embryonic stem cells. According to a further aspect of the invention there is provided a cell culture comprising: 10 primate embryonic stem cells on a proteinaceous based cell culture support and cell culture medium comprising: insulin, transferrin, sodium selenite, ethanolamine, 2 mercaptoethanol, oleic acid complexed with fatty acid free bovine albumin and further wherein the cell culture medium is supplemented with fibroblast growth factor and heparin characterised in that the cell culture further comprises at least one agent that 15 induces differentiation of the primate embryonic stem cells into at least one cell-type. In a preferred embodiment of the invention the primate embryonic stem cells are pluripotent human embryonic stem cells. 20 According to a further aspect of the invention there is provided a cell culture vessel comprising a cell culture medium that includes: insulin, transferrin, sodium selenite, ethanolamine, 2-mercaptoethanol, oleic acid complexed with fatty acid free bovine albumin and further wherein the cell culture medium is supplemented with fibroblast growth factor, heparin and ascorbic acid, or ascorbic acid phosphate, or derivative thereof. 25 In a preferred embodiment of the invention said cell culture vessel further comprises primate embryonic stem cells; preferably pluripotent human embryonic stem cells In a further preferred embodiment of the invention said vessel is selected from the group 30 consisting of: a petri-dish; cell culture bottle or flask; multiwell plate. "Vessel" is construed as any means suitable to contain a primate embryonic stem cell culture. 8 WO 2008/007082 PCT/GB2007/002584 According to a further aspect of the invention there is provided a cell culture medium container comprising a cell culture medium that includes: insulin, transferrin, sodium selenite, ethanolamine, 2-mercaptoethanol, oleic acid complexed with fatty acid free bovine albumin and further wherein the cell culture medium is supplemented with 5 fibroblast growth factor, heparin and ascorbic acid, or ascorbic acid phosphate, or derivative thereof. According to a further aspect of the invention there is provided a cell culture medium container comprising a cell culture media that includes: insulin, transferrin, sodium 10 selenite, ethanolamine, 2-mercaptoethanol, oleic acid complexed with fatty acid free bovine albumin and further wherein the cell culture medium is supplemented with fibroblast growth factor and heparin. Throughout the description and claims of this specification, the words "comprise" and 15 "contain" and variations of the words, for example "comprising" and "comprises", means "including but not limited to", and is not intended to (and does not) exclude other moieties, additives, components, integers or steps. Throughout the description and claims of this specification, the singular encompasses the 20 plural unless the context otherwise requires. In particular, where the indefinite article is used, the specification is to be understood as contemplating plurality as well as singularity, unless the context requires otherwise. Features, integers, characteristics, compounds, chemical moieties or groups described in 25 conjunction with a particular aspect, embodiment or example of the invention are to be understood to be applicable to any other aspect, embodiment or example described herein unless incompatible therewith. An embodiment of the invention will now be described by example only and with 30 reference to the following figures: Figure 1 illustrates the effect of bFGF on human embryonic stem cell proliferation; 9 WO 2008/007082 PCT/GB2007/002584 Figure 2 illustrates the effect of bFGF and heparin on human embryonic stem cell proliferation and morphology; Figure 3 illustrates the expression of human embryonic stem cell markers in cells cultured 5 in feeder free conditions; Figure 4 illustrates the growth of human embryonic stem cells in various medium; Figure 5 illustrates growth curves of human embryonic stem cell-line HUES in feeder free 10 conditions; and Figure 6 illustrates growth curves of human embryonic stem cell-line Shef 1 in feeder free conditions; and 15 Table 1 illustrates a summary of cell culture medium components for culturing human embryonic stem in feeder free conditions. Materials and MethodsFeeder Free Culture of Human Embryonic Stem Cells 20 hESF9 is defined in Table 1. Hesf5 medium is identical to Hesf9 medium without the addition of oleic acid complexed with bovine albumin, ascorbic acid phosphate, bFGF, and heparin sulphate. A. Reagents 25 1. T25 flask of human undifferentiated embryonic stem cells 2. hESF9 medium: ESF basal medium without HEPES supplemented with 9 factors, insulin, transferrin, sodium selenite, 2-mercaptoenthanol, 2-ethanolamine, oleic acid complexed with bovine albumin, ascorbic acid phosphate, bFGF, and heparin sulphate. 30 3. EDTA solution 4. Type I collagen (Nitta Gelatine, Co., Osaka, Japan) B. Procedure 1. Coat T25 (corning) with 100 Rg/cm 2 type I collagen. 10 WO 2008/007082 PCT/GB2007/002584 2. ES cell colonies were detached by 0.45mM to 0.5mM EDTA4Na (Sigma) in Dulbecco's phosphate buffered saline without Ca 2 and Mg 2 +. (The cells should not be dissociated into single cells. The concentration of EDTA depends on cell lines.) 3. Collect the cells by hESF9 medium. 5 4. Spin down the cell suspension for 3 min at 800 rpm. 5. Re-suspend the cells in hESF9 medium. 6. Spin down the cells suspension for 3 min at 800 rpm. 7. Re-suspend the cells in hESF9 medium. 8. The cells were seeded onto 25cm 2 flask coated with 100 [g/cm2 type I collagen in 10 hESF9 medium. 9. Incubate at 370 C in a humid atmosphere of 10% CO 2 . Neuronal Differentiation method A. Reagents 15 1. T25 flask of human undifferentiated embryonic stem cells 2. hESF9 medium 2. hESF5 medium: ESF basal medium without HEPES supplemented with 5 factors, insulin, transferrin, sodium selenite, 2-mercaptoenthanol and 2-ethanolamine. 3. EDTA solution 20 4. Laminin (sigma) 5. bFGF and heparin B. Procedure 1. Coat plastic dish by 5 jig/cm 2 laminin. 2. Undifferentiated ES cells are harvested by EDTA solution. 25 3. Seed the cells onto laminin-coated dish in hESF5 supplemented with lOng/ml bFGF and 100ng/ml heparin. Option. Seed the cells onto laminin-coated dish in hESF9 medium and on the next day, change the medium to hESF5 medium supplemented with 1Ong/ml bFGF and 1 OOng/ml heparin. 30 4. Culture at 370 C in a humid atmosphere of 5% CO 2 for one day. 5. On the next day, add 1Ong/ml bFGF into the culture. 6. On 2
~
4 th culture day, change the medium into hESF5 medium. 7. Every 2 days, change the medium to fresh hESF5 medium. 11 WO 2008/007082 PCT/GB2007/002584 8. On 7
~
1 0 th culture day, neuronal cells appear. Differentiation into epithelial-like cells. 5 A. Reagents 1. T25 flask of human undifferentiated embryonic stem cells 2. hESF9 medium 2. hESF5 medium supplemented with FA-BSA: ESF basal medium without HEPES supplemented with 5 factors, insulin, transferrin, sodium selenite, 2 10 mercaptoethanol, 2-ethanolamine, and 0.5 mg/ml fatty acid free bovine albumin (FA-BSA) 3. EDTA solution 4. Laminin (sigma) 5. BMP4 15 B. Procedure 1. Coat plastic dish by 5 jg/cm 2 laminin. 2. Undifferentiated ES cells are harvested by EDTA solution. 3. Seed the cells onto laminin-coated dish in hESF5 medium supplemented with FA-BSA and 1Ong/ml BMP4. 20 4. Every 2 days, change the medium to fresh hESF5 medium supplemented with FA-BSA and lOng/ml BMP4. 8. From 3 rd day of culture, epithelial-like cells appear. 25 30 12 WO 2008/007082 PCT/GB2007/002584 Table 1 Defined medium for feeder and serum free growth (hESF9). Components Concentration Components Concentration (mg/L) (mg/L) L-alanine 2.225 Vitamin B 1 2 0.34125 L-arginine 50 Hypoxanthine 1.02 L-arginine HCl 94.75 Oleic acid 9.4 L-asparagine H 2 0 16.2525 Linoleic acid 0.021 L-aspartatic acid 8.325 Lipoic acid 0.0525 L-cysteine HCl 7.88 Putrecine 2HCl 0.04025 L-cystine 2HCl 47.5725 Thymidine 0.1825 L-glutamatic acid 8.675 Sodium chloride 6599.75 L-glutamine 549.65 Potassium chloride 355.9 Glycine 19.375 Calcium chloride 108.305 (anhydrous) L-histidine 23.165 Calcium nitrate 4H 2 0 25 L-hydroxyproline 5 Magnesium chloride 14.305 (anhydrous) L-isoleucine 65.935 Magnesium sulfate 61.055 (anhydrous) L-leucine 68.225 Ferric nitrate 9H20 0.05 L-lysine HCI 92.175 Copper sulfate 5H 2 0 0.000625 L-methionine 19.87 Ferrous sulfate 7H20 0.2085 L-phenylalanine 37.99 Zinc sulfate 7120 0.216 L-proline 13.625 Sodium selenite 0.0034588 L-serine 31.125 Sodium dihydrogen 54.35 phosphate (anhydrous) L-threonine 55.525 Disodium 235.51 monohydrogen phosphate (anhydrous) L-tryptophan 9.76 Sodium pyruvate 110 L-tyrosine 42.36 2-mercaptoehanol 0.7813 L-valine 54.825 2-ethanolamine 0.6108 Glutathione 0.25 Insulin 10 Para-aminobenzoic acid 0.25 Apo-transferrin 5 Biotin 0.05185 Heparin sodium salt 0.1 Calcium pantothenate 2.1825 Albumine 1000 Choline chloride 6.24 Glucose (anhydrous) 2500 Folic acid 2.575 NaHCO 3 2000 Inositol 16.85 Phenol red 6.56 Niacinamide 2.25925 Fibroblast growth 0.01 factor-2 Pyridoxal HCI 2 Pyridoxine HCI 0.2655 Ascorbic acid phosphate 100 Riboflavin 0.2595 Thiamine HCI 2.335 5 13
Claims (48)
1. A method to maintain a primate embryonic stem cells in cell culture conditions that are 5 cell feeder and serum free comprising: forming a preparation of primate embryonic stem cells in a cell culture vessel comprising cell culture medium that includes fibroblast growth factor, heparin and ascorbic acid, or ascorbic acid phosphate, or derivative thereof and maintaining the primate embryonic stem cells in an undifferentiated state. 10
2. A method to maintain primate embryonic stem cells in cell culture conditions that are cell feeder free and serum free comprising the steps: i) forming a preparation of primate embryonic stem cells in a cell culture vessel which is coated with a proteinaceous based cell culture support wherein said cells are cultured in a cell culture medium comprising: insulin, transferrin, sodium selenite, ethanolamine, 2 15 mercaptoethanol, oleic acid complexed with fatty acid free bovine albumin and further wherein the cell culture medium is supplemented with fibroblast growth factor, heparin and ascorbic acid, or ascorbic acid phosphate, or derivative thereof; and ii) maintaining the primate embryonic stem cells in an undifferentiated state. 20
3. A method according to claim 1 or 2 wherein said primate embryonic stem cells have a stable karyotype.
4. A method according to any of claims 1-3 wherein ascorbic acid is ascorbic acid phosphate. 25
5. A method according to any of claims 1-4 wherein said primate embryonic stem cells are pluripotent human embryonic stem cells.
6. A method according to any of claims 1-5 wherein said primate embryonic stem cells 30 retain the property to differentiate into at least endoderm, mesoderm and ectoderm tissues throughout cell culture.
7. A method according to any of claims 1-6 wherein fibroblast growth factor (FGF) is selected from the group consisting of: aFGF, bFGF, FGF-4, FGF-9, FGF-17 or FGF- 18. 35
8. A method according to claim 7 wherein fibroblast growth factor is provided at a concentration of between 1-1 OOng/ml. 13 WO 2008/007082 PCT/GB2007/002584
9. A method according to claim 8 wherein fibroblast growth factor is provided at a concentration of about lOng/ml. 5
10. A method according to any of claims 1-9 wherein said fibroblast growth factor is bFGF.
11. A method according to any of claims 1-10 wherein fibroblast growth factor is recombinant. 10
12. A method according to any of claims 1-11 wherein ascorbic acid, or ascorbic acid phosphate, or derivative thereof is provided at a concentration of 0.01-0.2mg/ml.
13. A method according to claim 12 wherein ascorbic acid, or ascorbic acid phosphate, or derivative thereof is provided at about 0.1mg/ml. 15
14. A method according to any of claims 1-13 wherein 2-ethanolamine is provided at a concentration of 0.1-1 .0mg/ml.
15. A method according to claim 14 wherein 2-ethanolamine is provided at about 0.6mg/ml. 20
16. A method according to any of claims 1-15 wherein oleic acid is provided at a concentration of 3-15 gg /ml.
17. A method according to claim 16 wherein oleic acid is provided at about 9.5 pg /ml. 25
18. A method according to any of claims 1-17 wherein heparin is provided at a concentration of 10-500ng/ml. 30
19. A method according to claim 18 wherein heparin is provided at about 1OOng/ml.
20. A method according to claim 18 or 19 wherein heparin is heparin sodium salt.
21. A method according to any of claims 1-20 wherein said proteinaceous cell culture 35 support is collagen based. 14 WO 2008/007082 PCT/GB2007/002584
22. A method according to claim 21 wherein the collagen-based cell culture support comprises type I collagen.
23. A method according to claim 22 wherein type I collagen is recombinant type I collagen. 5
24. A method according to any of claims 1-20 wherein said cell culture support comprises recombinant human proteins selected from the group consisting of: collagen I, collagen IV, fibronectin, laminin and vitronectin. 10
25. A method according to claim 24 wherein said cell support comprises at least two recombinant proteins selected from the group consisting of: collagen IV, fibronectin, laminin and vitronectin.
26. A method according to claim 24 wherein said cell support comprises the recombinant 15 proteins collagen IV, fibronectin, laminin and vitronectin.
27. A method according to any of claims 1-20 wherein said proteinaceous cell culture support is Matrigelt". 20
28. A method according to any of claims 1-27 wherein said primate embryonic stem cells are passaged after addition of EDTA to the cell culture vessel.
29. A method according to any of claims 1-27 wherein said primate embryonic stem cells are passaged after addition of collagenase, preferably collagenase IV. 25
30. A method according to any of claims 1-27 wherein said primate embryonic stem cells are passaged after addition of dispase.
31. A method according to any of claims 1-27 wherein said primate embryonic stem cells are 30 passaged after addition of trypsin/EDTA, preferably recombinant trypsin.
32. A method according to any of claims 1-31 wherein said primate embryonic stem cells are cloned. 35
33. A method according to any of claims 1-32 wherein the cell culture media does not include the buffering agent HEPES. 15 WO 2008/007082 PCT/GB2007/002584
34. A method to differentiate primate embryonic stem cells into at least one cell-type in cell culture conditions that are cell feeder free and serum free comprising the steps: i) forming a preparation of primate embryonic stem cells in a cell culture vessel which is 5 coated with a proteinaceous based cell culture support wherein said cells are cultured in a cell culture medium comprising: insulin, transferrin, sodium selenite, ethanolamine, 2 mercaptoethanol, oleic acid complexed with fatty acid free bovine albumin and further wherein the cell culture medium is supplemented with fibroblast growth factor and heparin; and ii) adding an agent that induces the differentiation of the primate embryonic stem cells into 10 at least one cell-type.
35. A method according to claim 34 wherein the primate embryonic stem cells are human pluripotent embryonic stem cells. 15
36. A method according to claim 34 or 35 wherein the cell-type is a neurone.
37. A method according to claim 34 or 35 wherein the cell-type is an epithelial cell.
38. A method according to any of claims 34-37 wherein said proteinaceous based cell culture 20 support is laminin.
39. A cell culture comprising: primate embryonic stem cells on a proteinaceous based cell culture support and cell culture media comprising: insulin, transferrin, sodium selenite, ethanolamine, 2-mercaptoethanol, oleic acid complexed with fatty acid free bovine albumin and 25 further wherein the cell culture medium is supplemented with fibroblast growth factor, heparin and ascorbic acid, or ascorbic acid phosphate, or derivative thereof.
40. A cell culture according to claim 39 wherein the primate embryonic stem cells are pluripotent human embryonic stem cells. 30
41. A cell culture comprising: primate embryonic stem cells on a proteinaceous based cell culture support and cell culture medium comprising: insulin, transferrin, sodium selenite, ethanolamine, 2-mercaptoethanol, oleic acid complexed with fatty acid free bovine albumin and further wherein the cell culture medium is supplemented with fibroblast growth factor and 35 heparin which cell culture further comprises at least one agent that induces differentiation of the primate embryonic stem cells into at least one cell-type. 16 WO 2008/007082 PCT/GB2007/002584
42. A cell culture according to claim 41 wherein the primate embryonic stem cells are pluripotent human embryonic stem cells. 5
43. A cell culture vessel comprising a cell culture medium that includes: insulin, transferrin, sodium selenite, ethanolamine, 2-mercaptoethanol, oleic acid complexed with fatty acid free bovine albumin and further wherein the cell culture medium is supplemented with fibroblast growth factor, heparin and ascorbic acid, or ascorbic acid phosphate, or derivative thereof. 10
44. A cell culture vessel according to claim 43 wherein said cell culture vessel further comprises primate embryonic stem cells.
45. A cell culture vessel according to claim 43 or 44 wherein said primate embryonic stem cells are pluripotent human embryonic stem cells. 15
46. A cell culture vessel according to any of claims 43-45 wherein said vessel is selected from the group consisting of: a petri-dish; cell culture bottle or flask; multiwell plate.
47. A cell culture medium container comprising a cell culture medium that includes: insulin, 20 transferrin, sodium selenite, ethanolamine, 2-mercaptoethanol, oleic acid complexed with fatty acid free bovine albumin and further wherein the cell culture medium is supplemented with fibroblast growth factor, heparin and ascorbic acid, or ascorbic acid phosphate, or derivative thereof.. 25
48. A cell culture medium container comprising a cell culture media that includes: insulin, transferrin, sodium selenite, ethanolamine, 2-mercaptoethanol, oleic acid complexed with fatty acid free bovine albumin and further wherein the cell culture medium is supplemented with fibroblast growth factor and heparin. 30 35 17
Applications Claiming Priority (3)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| GBGB0613756.6A GB0613756D0 (en) | 2006-07-12 | 2006-07-12 | Cell culture medium |
| GB0613756.6 | 2006-07-12 | ||
| PCT/GB2007/002584 WO2008007082A2 (en) | 2006-07-12 | 2007-07-10 | Cell growth medium |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| AU2007274065A1 true AU2007274065A1 (en) | 2008-01-17 |
Family
ID=36955441
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| AU2007274065A Abandoned AU2007274065A1 (en) | 2006-07-12 | 2007-07-10 | Cell growth medium |
Country Status (7)
| Country | Link |
|---|---|
| EP (1) | EP2038404A2 (en) |
| JP (1) | JP5227318B2 (en) |
| CN (1) | CN101490243A (en) |
| AU (1) | AU2007274065A1 (en) |
| CA (1) | CA2657539A1 (en) |
| GB (2) | GB0613756D0 (en) |
| WO (1) | WO2008007082A2 (en) |
Families Citing this family (33)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP1962719A4 (en) | 2005-08-29 | 2011-05-04 | Technion Res And Dev Of Foundation Ltd | Media for culturing stem cells |
| DK3441459T3 (en) | 2006-08-02 | 2021-06-07 | Technion Res & Dev Foundation | PROCEDURES FOR EXPANSION OF EMBRYONAL STEM CELLS IN A SUSPENSION CULTURE |
| US9688956B2 (en) | 2008-06-05 | 2017-06-27 | National Cheng Kung University | Method for preserving proliferation and differentiation potential of mesenchymal stem cells |
| CN102076280B (en) | 2008-06-24 | 2014-08-27 | 生物活性外科公司 | Surgical sutures incorporated with stem cells or other bioactive materials |
| KR20110042107A (en) | 2008-08-07 | 2011-04-22 | 바이오엑티브 써지컬, 아이엔씨. | Stem cell capture and immobilization coatings for medical devices and implants |
| CN102317442B (en) * | 2008-12-17 | 2014-08-13 | 斯克里普斯研究所 | Generation and maintenance of stem cells |
| TW201109440A (en) * | 2009-07-21 | 2011-03-16 | Transgene Sa | Enzymatic composition for the digestion of chicken embryos |
| JP6276918B2 (en) | 2009-11-12 | 2018-02-07 | テクニオン リサーチ アンド ディベロップメント ファウンデーション リミテッド | Medium, cell culture and method for culturing pluripotent stem cells in undifferentiated state |
| CN101812480B (en) * | 2009-11-16 | 2013-04-10 | 西北农林科技大学 | Method utilizing transcription factor to transfect bovine body cell into induced pluripotent stem cell |
| JP5714267B2 (en) * | 2010-08-26 | 2015-05-07 | 日本メナード化粧品株式会社 | Stem cell undifferentiation maintenance agent and proliferation promoter |
| JP6148429B2 (en) * | 2011-01-31 | 2017-06-14 | 協和発酵バイオ株式会社 | Method for culturing human pluripotent stem cells |
| CN102586176B (en) * | 2012-01-11 | 2013-05-08 | 中国科学院生物物理研究所 | Novel animal source-free and feed layer-free human pluripotent stem cell culture system |
| GB201211873D0 (en) * | 2012-07-04 | 2012-08-15 | Univ Edinburgh | Cell culture |
| SG11201505965TA (en) * | 2013-01-31 | 2015-09-29 | Ajinomoto Kk | Culture method for stable undifferentiated proliferation of pluripotent stem cells |
| SG11201509767UA (en) * | 2013-05-30 | 2015-12-30 | Ajinomoto Kk | Medium for culturing stem cells |
| CN105358707B (en) * | 2013-06-28 | 2019-06-14 | 株式会社钟化 | The method for screening multipotential stem cell positive growth factor |
| WO2015042356A1 (en) * | 2013-09-19 | 2015-03-26 | The United States Of America, As Represented By The Secretary, Department Of Health & Human Services | Chemically defined culture medium for stem cell maintenance and differentiation |
| CN104357379B (en) * | 2014-09-30 | 2017-08-08 | 刘兴宇 | Stem cell media |
| CN106032527B (en) * | 2015-03-17 | 2019-08-13 | 广州市搏克肿瘤研究所 | It is a kind of tolerance low-density without feeder layer human pluripotent stem cells culture medium |
| CN105907705B (en) * | 2016-06-28 | 2020-01-24 | 广州市搏克生物技术有限公司 | Pluripotent stem cell culture medium |
| JP7049347B2 (en) * | 2017-01-23 | 2022-04-06 | ステムセル テクノロジーズ カナダ インコーポレイテッド | Mediums and Methods to Enhance Stem Cell Survival and Proliferation |
| CN106754715A (en) * | 2017-02-13 | 2017-05-31 | 四川新生命干细胞科技股份有限公司 | A kind of trophoblastic preparation method for candidate stem cell culture |
| CN106754652B (en) * | 2017-03-06 | 2019-04-02 | 广州润虹医药科技股份有限公司 | IPS cell differentiation at ectoderm progenitor cells serum-free induced medium and abductive approach |
| CN106754657B (en) * | 2017-03-28 | 2022-07-22 | 北京赛斯达生物技术有限公司 | Serum-free medium for monkey embryonic stem cells |
| ES2955861T3 (en) * | 2017-05-31 | 2023-12-07 | Promocell Gmbh | Culture medium for pluripotent stem cells |
| CN110157662A (en) * | 2018-02-15 | 2019-08-23 | 郭永珍 | A kind of cell additive and preparation method thereof |
| KR102147780B1 (en) * | 2018-03-16 | 2020-08-25 | 사회복지법인 삼성생명공익재단 | Culture medium composition for promoting the proliferation of stem cell comprising FGF-17 as an effective ingredient and method of culturing stem cells using the same |
| CA3104838A1 (en) * | 2018-06-27 | 2020-01-02 | Ajinomoto Co., Inc. | Additive for culturing stem cells, culturing medium, and culturing method |
| CN110684716A (en) * | 2018-07-08 | 2020-01-14 | 郭永珍 | Preparation method of cell additive and product thereof |
| CN109486766B (en) * | 2018-11-26 | 2021-05-07 | 中山大学 | Lacrimal gland stem cell, culture system and culture method of lacrimal gland stem cell |
| CN111484970B (en) * | 2020-04-30 | 2022-09-16 | 广州再生医学与健康广东省实验室 | Serum-free and feeder-layer-free embryo and pluripotent stem cell culture medium with low protein content |
| WO2023150293A2 (en) * | 2022-02-03 | 2023-08-10 | Steakholder Foods Ltd. | Accelerated myotube formation |
| CN116210647A (en) * | 2023-02-03 | 2023-06-06 | 威奥福生物科技(宁波)有限公司 | Method for propagating white-feather broiler chickens by using primordial germ cell stem cell line |
Family Cites Families (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| AU5734998A (en) * | 1997-01-10 | 1998-08-03 | Life Technologies, Inc. | Embryonic stem cell serum replacement |
| US7439064B2 (en) * | 2000-03-09 | 2008-10-21 | Wicell Research Institute, Inc. | Cultivation of human embryonic stem cells in the absence of feeder cells or without conditioned medium |
| WO2003029443A1 (en) * | 2001-09-28 | 2003-04-10 | Es Cell International Pte Ltd | Methods of derivation and propagation of undifferentiated human embryonic stem (hes) cells on feeder-free matrices and human feeder layers |
| CA2527847C (en) * | 2003-06-12 | 2015-09-29 | Yeda Research & Development Co. Ltd | Enhancement of oligodendrocyte differentiation |
| EP1698690B1 (en) * | 2003-12-26 | 2010-04-28 | Makoto Asashima | Basal medium for es cell culturing |
| GB2431165B (en) * | 2004-07-13 | 2009-04-01 | Geron Corp | Medium for growing human embryonic stem cells |
| GB2432846B (en) * | 2004-09-08 | 2009-12-30 | Wisconsin Alumni Res Found | Medium and culture of embryonic stem cells |
-
2006
- 2006-07-12 GB GBGB0613756.6A patent/GB0613756D0/en not_active Ceased
-
2007
- 2007-07-10 WO PCT/GB2007/002584 patent/WO2008007082A2/en active Application Filing
- 2007-07-10 CA CA002657539A patent/CA2657539A1/en not_active Abandoned
- 2007-07-10 JP JP2009518958A patent/JP5227318B2/en active Active
- 2007-07-10 EP EP07766180A patent/EP2038404A2/en not_active Withdrawn
- 2007-07-10 AU AU2007274065A patent/AU2007274065A1/en not_active Abandoned
- 2007-07-10 CN CNA2007800249934A patent/CN101490243A/en active Pending
-
2008
- 2008-12-29 GB GB0823575A patent/GB2452456A/en not_active Withdrawn
Also Published As
| Publication number | Publication date |
|---|---|
| CN101490243A (en) | 2009-07-22 |
| CA2657539A1 (en) | 2008-01-17 |
| WO2008007082A2 (en) | 2008-01-17 |
| JP2009542247A (en) | 2009-12-03 |
| JP5227318B2 (en) | 2013-07-03 |
| GB0613756D0 (en) | 2006-08-23 |
| GB0823575D0 (en) | 2009-01-28 |
| EP2038404A2 (en) | 2009-03-25 |
| WO2008007082A3 (en) | 2008-03-06 |
| GB2452456A (en) | 2009-03-04 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| AU2007274065A1 (en) | Cell growth medium | |
| US10047341B2 (en) | Generation of keratinocytes from pluripotent stem cells and maintenance of keratinocyte cultures | |
| AU2005282414B2 (en) | Culturing human embryonic stem cells | |
| CA2807418C (en) | Simplified basic media for human pluripotent cell culture | |
| AU2011358083B2 (en) | Method for culturing human pluripotent stem cells | |
| US20070010011A1 (en) | Defined media for pluripotent stem cell culture | |
| US20080124801A1 (en) | Pluripotent cell growth media | |
| MX2007002390A (en) | Medium and culture of embryonic stem cells. | |
| US20120164729A1 (en) | Composition for culturing pluripotent stem cells and use thereof | |
| JP6913694B2 (en) | Culture medium for pluripotent stem cells | |
| IL263557A (en) | Differentiation of pluripotent stem cells into corneal cells | |
| CA2504179A1 (en) | Composition for culturing multipotent stem cells and utilization of the same | |
| IL292933A (en) | Serum-free human pluripotent stem cell culture medium | |
| Ludwig et al. | Defined culture media for human embryonic stem cells |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| MK1 | Application lapsed section 142(2)(a) - no request for examination in relevant period |