Protein measurements in 2% Triton X samples
Posted 11 January 2012 - 08:22 AM
do any of you know what techniques can be used to determine protein quantity in cell lysates that contain 2% triton x. I have lysed my cells in lysis buffer containing 2% triton, because I am looking at a membrane bound protein (LC3) by WB.
Bradford etc. reagents are reacting very! good with Triton X, and the reagent gets saturated very fast by Triton.
Posted 12 January 2012 - 08:22 AM
Edited by mdfenko, 12 January 2012 - 08:24 AM.
genius does what it must
i do what i get paid to do
Posted 12 November 2013 - 07:37 PM
Because triton reacts with coomassie blue (Bradford reagent), I ran a serial dilution of triton x-100 while keeping the rest of the buffer the same (no protein). The serial dilution was the following:
2%, 1%, 0.5%, 0.25%, 0.125%, 0.0625%, 0.03125%, 0.0156%, and 0% triton x-100 (in HBSS).
I found that loss of interference by triton x occurred somewhere near 0.0625%, which could explain why many lysis buffer recipes only use 0.05% triton x (presumably because their designers were using the Bradford assay)
Thus if you find sufficient yield with as low ad 0.05% triton, the Bradford is a suitable quantification technique.
Edited by JeremyDNA, 12 November 2013 - 09:09 PM.