作者eriel (橘子数)
看板Biotech
标题[问题]关於免疫萤光法
时间Mon Nov 6 14:45:33 2006
我是参考Cell Signaling Technology 03-04
里面第396页提到
Cell fixation有两种方法:
1.Methnol (protein precipitation) fixation:
Immerse coverslips in 100% methnol at -20C for 10 min
2.Paraformaldehyde (cross-linking) fixation:
2a Immerse coverslips in 4% paraformaldehyde at room temp. for 10 min
2b Wash coverslips once with PBS
2c Aspirate completely and then permeabilize cells with 0.2% Triton X-100
for 5 min at room temp. or, alternatively with methnol at -20C for 5 min.
请问这两种fixation方式究竟有何不一样？(cross-linking and protein precipitation)
还有我的sample是sperm ，想要染出膜上或膜内的某个蛋白domain，我该fixation吗?
最後上述2C，paraformaldehyde後，改以triton或methnol有何不同呢？
问题很多，麻烦大家，谢谢
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