从我开始接触培养细胞已经有将近两年的时间，从最开始的细胞半死不活，到现在一个人同时培养多种细胞不出乱子，其中的每一点改变都是被老板骂的狗血淋头之后的悔悟。（因为本人主要接触到的细胞培养主要是人或小鼠的正常组织或癌组织细胞的已经被命名细胞系或细胞株的细胞，所以我下面所讲的具体方法也主要针对于此。如有不同见解，欢迎一起探讨）
培养细胞可以说是一个需要花费大量时间和大量精力的过程，如果你有一个有钱的老板还好，至少你不用为了清洗瓶瓶罐罐而痛苦。如果没有，就自求多福吧~。
在细胞培养方面我们主要需要注意的几个方面分别是：
- 你所培养的细胞所需要满足的环境条件和一般细胞有没有区别（37℃，5%CO2）
- 你的细胞所使用的血清是哪一种
- 你的细胞所使用的基础培养基是哪一种
- 给你该细胞的人在传代冻存时的一些细节
- 当然最最重要的一点是保证自己的操作不会污染细胞！！！
所以这次我们的主题就是养细胞之前需要注意什么。下面我就从保证自己不会污染细胞，开始给大家具体讲一下：
我们把培养细胞的地方叫做细胞间，一般需要达到二级生物安全房（P2）的标准。我们首先要保证环境的相对无菌，然后是人员和所带入物品的无菌最后才是操作。细胞间因为本身实验的要求需要有彻底的大扫除（半年或一学期），深层次清洁（每周），日常清洁（每天）和每次实验（需要打扫好自己的实验区域）。这些内容会在之后的文章中给出。
进入细胞间之前，先打开紫外灯（房间的），拿好所有要用的试剂（应该是无菌的，然后用封口条封住的），容器（废液缸）等物品。这里需要说一下，有一些试剂是不能高压灭菌的，需实验前在细胞间内单独用0.22um的滤器过滤（注意溶液的脂溶性还是水溶性，要和过滤器的类型对应）。DMSO本身无菌，不能高压也不能过滤。（建议细胞间的DMSO使用SGMA的）
等待紫外灯工作半个小时以后，关闭紫外灯，等待5分钟后进入缓冲间，更换拖鞋，穿实验服（建议手术服或全包安全服）戴帽子，口罩，全身喷70%酒精，最后戴手套再喷70%酒精进入细胞间。
所携带的物品需高压灭菌后带入，有无菌外包装的可以喷70%酒精后带入。
打开水浴锅（37℃），放入需要复温的培养基，缓冲溶液，胰蛋白酶等。
打开生物安全柜或超净工作台的紫外灯。
喷酒精然后擦拭倒置显微镜。
从CO2培养箱中取出细胞，在显微镜下观察细胞形态，细胞密度，是否有污染等。
等待超净工作台或生物安全柜的紫外灯打开15分钟后，先打开风机（不要过大）后推开玻璃面板，最后开灯。手进入里面时必须喷70%酒精（手臂处也要喷）。70%酒精擦拭工作台你实际操作的部分，点燃酒精灯。就可准备移入细胞，培养基等。
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