[求救] SDS-PAGE跑出來的protein大小與MS結果不符合?
各位好,
最近在做SDS-PAGE(12%,sample有加2-ME)
由於想要鑑定表現量有差異的protein,
所以之後以LC-MS/MS(orbitrap)作分析
因為是一維的band所以一定不止只有一種protein.
但是結果讓我很困惑...
如果是切50k的band但是有對到一些比50k大的protein應該算正常,
因為可能是一些subunit,或是打斷雙硫鍵的片段.
但比如我是切100k的band去鑑定,但結果卻有對到30K的protein,或是70k的albumin等等
如果denature沒完全有差那麼多嗎?modification應該也不是這樣阿?
甚至有些band連質量差不多相符的protein都沒有..不是比切的位置大要不就是小很多
但MS靈敏度高,的確是證實有這些protein...(已去除false positive)!
謝謝各位看完~~
請各位高手解惑~~感謝!
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