Is mCherry denatured by Trypsin or RIPA buffer?
Posted 01 September 2011 - 04:38 PM
I am currently transfecting cells with mCherry and lysing the cells to determine the fluorescence of mCherry. However, it has just been brought to my attention that using trypsin to lift my cells and/or my lysis buffer may affect my mCherry protein. I can't find anything in the literature about it, so I was hoping someone may have some knowledge in this? Thanks!!!
Posted 01 September 2011 - 08:09 PM
Couldn't you determine the fluorescence on cells on a slide?
Posted 05 September 2011 - 05:31 AM
Posted 06 September 2011 - 12:53 PM
Is it possible to determine the fluorescence on a slide after I have lysed the cells?
No I am not using protease inhibitors. Do you mean I should use protease inhibitors when I'm lysing my cells with the Ripa buffer?
Posted 06 September 2011 - 03:20 PM
You should always use protease inhibitors when lysing, this is to stop your proteins from degrading quite so quickly! Unless you have washed the cells very well there will still be some trypsin with the cells after lifting, and they also contain native proteases.
Posted 08 September 2011 - 08:43 AM
Posted 08 September 2011 - 09:54 PM
Posted 14 September 2011 - 10:47 AM