bands from tissue one size, from cells - size doubles
Posted 03 February 2005 - 09:56 PM
From mouse tissues I see no other bands appearing other than a strong band at 30kDa, but with the cell lysates I see a faint shadow at 30kDa but a very strong band at 60kDa.
A colleague has suggested that maybe my protein has formed dimers, strongly linked that they are not dissociated in the reducing sample buffer. Could this be the case, or does anyone have any other ideas???
Thankyou in advance,
Posted 04 February 2005 - 09:29 AM
I guess its the dimer of your protein. What concentration of reducing agent you are using in your loading buffer? first of all is it mercapto ethanol or DTT? and also some times boiling the samples before loading helps. did u try that?
Posted 06 February 2005 - 05:47 PM
Thanks for your reply,
Posted 07 February 2005 - 01:20 PM
Posted 07 February 2005 - 09:28 PM
i have been using this buffer on my cells as others in the lab are using it successfully to view phosphorylated proteins, which is one thing I want to see from my cells. I guess the trizol and homogenisation could be breaking apart any dimers.
How can I tell for sure? How do you deternmine if a protein is dimerising???