b9602015 chia-chen li 1 el pilar. 2 what is cell culture?! (1) maintaining cell line survival under...
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IFMSA-Zaragoza Project
B9602015Chia-Chen Li
El Pilar
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Cell Culture
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What is CELL CULTURE?! (1)
• Maintaining cell line survival under controlled conditions, in despite of their original environment
– Proliferative capacity– Differentiation ability– Corresponding cell age
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What is CELL CULTURE?! (2)
• Controlled conditions– ex vivo– in vitro
• Tissue source– Multi-cellular eukaryotes– Mainly animals– Also plants, fungi & microbes
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General Requirements (1)
• Sterilization – Methanol– Filtering by 0.20 μm
• Culture surfaces – Petri dish, flake or plate
• Incubation – Temperature at 37℃
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General Requirements (2)
• Nutrient media – Antibiotic– Buffering• efficient at pH 7.2-7.6
– CO2/pH detectable• Red (pH 8, fresh)
Yellow (pH 6, wastes)
• Trypsin, the growth inhibitor
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Daily Care
• Changing media (yellow)• Microscopy observation for cell growth
monitoring, monolayer • Trypsin– Reduction for multilayer culture– Dividing for new flakes
• Photographing
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Results (1)
• Breast tumor – derived from nude mice
• Hyperchromatic• Hyper-proliferative
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Prominent nuclei
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Results (2)
• Breast tumor – from nude mice
• Duct• Ductules
duct
ductules
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Results (3)
• Breast tumor – derived from
nude mice
• Epithelial cell alignment
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cell alignment
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Animal Experiment
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Athymic Nude Mice
• Spontaneous genetic mutated mice– FOXN1 deletion
• Immunodeficient– Lack of mature T cell– Lack of adaptive
immune response
• Xenograft transplant – Human prostate tumor
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Woking Conditions
• Requirements – Working robe– Gloves – Shoe pads
• Not allowed– Bare-handed
touching– Sniffing & etc.
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Daily Care for Nude Mice
• Provides– Meal, sterilized – Water, sterilized – Bed surface
• Used cages must be washed and sterilized for another-time use.
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Experiment Design
• Intra-tumor injection, 5 times / 3 weeks– Control
• normal saline buffer
– Variables • Anti-proliferative ligands• Granulysin
• Tumor size measuring– Every injection – The middle of each injection intervals
Under anaesthesia
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Efficiency Determination
• By square a*b, the 2D area of tumor
• Effective potency:
• The effectiveness can be confirmed if there’s significant change of the potency, in which the tumor size reduces.Experiment is still in progress.
ab
a1*b1 - a0*b0
a0*b0
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Thanks!
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