Protocol Online logo  Protein extraction - (May/17/2012 )   I am
trying to extract proteins from MCF7 and BT20 cell lines in
order to analyze through a western blot. The protein of interest
is Cyp1b1 which is Flag tagged. Since Cyp1b1 is bound to the
smooth ER, I purchased a Pierce Protein Extraction Kit
(http://www.piercenet...CFQSynQodDRNGLw).  Since I am going to
be looking at protein degradation at different time points
(cyclohexamide treatment and protein extracted 0hrs, 4hrs, and
8hrs), I was told that I would need protease inhibitors to keep
my protein from being degraded after extraction. This is the
first time my lab has worked with human cell lines, and nobody
is 100% sure of what all that entails. I hope somebody on here
can help me out a bit as to what I need to purchase in order to
extract the protein which will be used for westerns.  Thanks in
advance,   Typically you would make what is known as a lysate -
this is basically dissolved cells in a solution that contains
detergent(s) (often NP-40, n