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<div style="color: black !important; font-family: 'Times New Roman', SimSun, serif !important, line-height:1.5;"> == 细胞总蛋白提取 == *'''配液:细胞裂解液=PIRP:PMSF=100:1, Loading buffer 蛋白上样缓冲液(5×), β-巯基乙醇(保护巯基)''' *'''提取:1.六孔板PBS清洗,消化,离心,移至1.5ml离心管内,PBS洗一次 2.放置于冰盒上,每孔加裂解液50-60 ul,吹打均匀,4℃裂解30 min 3. 4℃,12000-14000 rpm,15 min离心 4. 取上清液于0.5 ml离心管内,每次吸取10 ul定量 5.取1-2 ul用作BCA检测,剩余按上清液:Loading buffer=4:1 == 组织蛋白提取 == *'''提取:把组织剪成细小碎片,复溶细胞裂解液,按每20 mg组织加入100-200 ul裂解液,用玻璃匀浆器匀浆,直至充分裂解,4℃ 10000-14000离心10-15 min,取上清液,取样进行BCA检测,剩余4:1与蛋白上样缓冲液混匀,煮沸5-10 min,分装-80℃保存(45 ul/管,一次用20 ul上样,每孔一般30-40 ug,上样体积10-15 ul)
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