国外蛋白组学专业BBS的一些问答-强烈推荐（11-14更新）
丁香园论坛
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1,关于使用三氯醋酸沉淀法后蛋白质难于重悬的问题
Q:1,Hi !I have been trying to clean and concentrate proteins form eucariotic cell culture medium in order to analyse them in 2D. I’ve read some papers were TCA precipitation was used and recommended for this purpose but I have not been able to redissolve my samples. I have used 10%TCA and let it precipitate for 30 minutes in Ice, afterwards I have centrifuge the sample, washed it with diethil ether and tried to dissolve the pellet in ordinary IEF sample solution containing Urea, Chaps, ampholites and DTT. I wasn’t able to dissolve the sample even using a 9M Urea concentration.
Do you have any suggestions on how to dissolve proteins from TCA precipitation or an alternative way to clean and concentrate proteins from culture medium for 2d analysis?
Thank you!
A:This problem occurs very often.
Better for cleaning and concentration of protein samples: Use the Ettan 2-D Cleanup Kit from Amersham Biosciences
codegreen点评:这个问题由AMERSHAM著名科学家回答,比较会替公司做广告.
其实真核细胞总蛋白质的提取用不着使用TCA/acetone法,细胞相对于植物组织还是比较干净的,少掉了一些脂类\酚类杂质.对于组织来讲,使用clean-up kit效果还是不错的,等过几天做实验了,小弟把自己做的使用clean-up kit前后的2-D图谱之对比贴出来.
Q:1,Hi !I have been trying to clean and concentrate proteins form eucariotic cell culture medium in order to analyse them in 2D. I’ve read some papers were TCA precipitation was used and recommended for this purpose but I have not been able to redissolve my samples. I have used 10%TCA and let it precipitate for 30 minutes in Ice, afterwards I have centrifuge the sample, washed it with diethil ether and tried to dissolve the pellet in ordinary IEF sample solution containing Urea, Chaps, ampholites and DTT. I wasn’t able to dissolve the sample even using a 9M Urea concentration.
Do you have any suggestions on how to dissolve proteins from TCA precipitation or an alternative way to clean and concentrate proteins from culture medium for 2d analysis?
Thank you!
A:This problem occurs very often.
Better for cleaning and concentration of protein samples: Use the Ettan 2-D Cleanup Kit from Amersham Biosciences
codegreen点评:这个问题由AMERSHAM著名科学家回答,比较会替公司做广告.
其实真核细胞总蛋白质的提取用不着使用TCA/acetone法,细胞相对于植物组织还是比较干净的,少掉了一些脂类\酚类杂质.对于组织来讲,使用clean-up kit效果还是不错的,等过几天做实验了,小弟把自己做的使用clean-up kit前后的2-D图谱之对比贴出来.
