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Material
Sacrifice Animals Place mice in a bag and fill bag with CO2 - 3-5 minutes, alternatively Dry ice method Pour Betadine over head Spray animals with Etoh Cut Skin sagitally from back of neck towards nose Remove calvaria with scissors-place in PBS on ice in a petri dish Scrape soft tissues off calvaria macroscopically and microscopically (total of 2 times) Place in PBS Mince with scissors into small pieces Wash 2-3× in successive dilutions of Betadine:PBS and finally with PBS Place in Digestive media in a conical tube (75 ml DMEM -alpha , 0.150g Dispase II, 0.075 g Collagenase A) Shake at 37℃ for 10 minutes - Discard first round of digestion - too many fibroblasts and unwanted cells - collect the subsequent supernatants Wash and spin (1000 rpm) 3× with PBS to remove enzymes Plate in Media with serum (10%) |
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上一篇细胞培养方法的问答（1） 下一篇 电穿孔和电融合技术
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