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人皮肤成纤维细胞CCC-HSF-1一、细胞操作参考书
二、生长特性:贴壁 悬浮
三、细胞生长条件:
培养基 90% (GIBCO)
血清 10% FBS(GIBCO)
温度 37℃
空气条件 5% CO2,,95% AIR
生长代数 P4
冻存条件 培养基70%、血清20%、DMSO10%
四、组成:
组份 规格
细胞一瓶 T25
细胞培养与操作说明 1份
五、细胞接收后的操作流程与注意事项
1.如果细胞为贴壁细胞,,而收到时呈悬浮或者部分悬浮的状态,请将悬浮的细胞即时离心,加15%血清的*培养基到新的培养皿/瓶继续培养3天;同时原培养瓶中剩下的贴壁细胞更换为15%血清的*培养基,培养3天。3天后若原瓶或者新瓶中的细胞都没有出现增殖而是继续脱落死亡,请及时实验室,技术人员会跟进解决
2.贴壁细胞生长缓慢:适当提高血清浓度(zui高不能超过20%),或可根据该细胞生长密度,考虑胰酶消化后,转移到新的培养瓶继续培养
3.生长不均:贴壁细胞若出现分布不均,成岛状生长,可将细胞进行消化,重悬打散细胞,加入新鲜培养基进行培养
六、细胞常规培养传代流程(请严格遵照无菌操作)
1.吸出原培养瓶中的培养基,PBS缓冲液润洗细胞两次,加2~3 ml 0.25%胰酶进行消化细胞(注意把握消化时间,通常控制在1-2min)
2.镜下观察消化情况,在细胞边缘缩小,贴壁松动时(不建议消化到细胞漂浮)去掉胰酶,加6~8ml *培养基,轻轻吹打细胞层,尽量把细胞层吹落,吹散
3.取部分细胞悬液转移到新的培养皿/瓶中,添加适当的*培养基,,把细胞悬液打匀,于培养箱中培养
4.注意培养基PH值变化情况,定期换液(每周2-3次),待细胞密度达到80%以后重复1项操作或者冻存
特别注意:(如使用公共实验室或初次接触细胞培养,建议添加双抗培养)
1.收到细胞后请尽快更换为含15%血清的新鲜培养基,如因特殊情况需要继续使用原瓶培养基,请在原瓶培养基中额外添加10%的血清(原瓶培养基的继续使用时间zui长不宜超过72小时)
2.贴壁细胞收到当天切忌立刻消化,请将细胞放置培养箱孵育过夜到第二天再做消化传代
如签收时出现培养瓶壁破裂,漏液等情况请及时做好照片记录并实验室
TNS-sf (Trypsin Neutralization Solution-serum free)
Enzyme-Free Cell Dissociation Solution
Cell Freezing Medium
Serum-Free Cell Freezing Medium
Human Pluripotent Stem Cell Cryopreservation Medium
Human Pluripotent Stem Cell Cryopreservation Medium,
Trypan Blue, 0.4%
Dulbecco’s Phosphate-Buffered Saline
Hank's Balanced Salt Solution
Poly-L-Lysine 1 mg/ml
Poly-L-Lysine 10 mg/ml
0.2% Gelatin Solution
Fetal Bovine Serum, Human Cell Tested
Penicillin/Streptomycin Solution
Penicillin/Streptomycin Solution
Antimycotic Solution
Antibiotic/Antimycotic Solution
Cell Culture Grade Water
Myocyte Detachment Solution
Bovine Pituitary Extract
Bovine Pituitary Extract
Insulin-Transferrin-Selenium, 100x
L-Glutamine Solution, 200 mM
100X Non-Essential Amino Acids
Human Germ Layer Detection Kit
Oil Red O Staining Kit
StemDS? Human Embryonic Stem Cell Dissociation Solution
Bovine Plasma Fibronectin
Human Cellular Fibronectin
Bovine Plasma Vitronectin
"Recombinant Human Serum Albumin
"
Bovine Plasma Fibronectin
Collage I-Cell Culture Surface Coating Kit
Poly-L-Lysine 1 mg/ml
Poly-L-Lysine 10 mg/ml
Human Pluripotent Stem Cell Growth Medium
Xeno-free Human Pluripotent Stem Cell Growth Medium
Animal Component-free Human Pluripotent Stem Cell Growth Medium
StemDS? Human Embryonic Stem Cell Dissociation Solution
Human Pluripotent Stem Cell Cryopreservation Medium
Human Pluripotent Stem Cell Cryopreservation Medium,
Mouse Embryonic Fibroblasts
Mouse Embryonic Fibroblasts-mitomicine treated
Mouse Embryonic Fibroblasts- Mitomycin C treated from CD1 mouse
Mouse Embryonic Fibroblasts from CF1 mouse
Mouse Embryonic Fibroblasts-Mitomycin C treated from CF1 mouse
Human Dermal Fibroblasts-Fetal-Mitomycin C treated
Mouse Embryonic Fibroblasts-Conditioned Medium
bFGF-Stimulated Mouse Embryonic Fibroblasts Conditioned Medium
Ultra-Low Binding Culture Plate
Ultra-Low Binding Culture Plate (6-well)
Ultra-Low Binding Culture Plate (24-well)
Ultra-Low Binding Culture Plate (24-well)
Recombinant Human Fibroblast Growth Factor-basic
"Recombinant Human Epidermal Growth Factor
"
"Recombinant Human Bone Morphogenetic Protein-2
"
"Recombinant Human Bone Morphogenetic Protein-4
"
"Recombinant Human Bone Morphogenetic Protein-7
"
"Recombinant Human NOGGIN
"
Human Amniotic Mesenchymal Stromal Cells
Human Chorionic Mesenchymal Stromal Cells
Human Mesenchymal Stem Cells-bone marrow
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