protein sample floating out of the tricine gel - (Jan/15/2013 )

I am writting to ask whether anybody had a same problem as mine. My protein samples are floating out of the Tricine-SDS-PAGE. I prepared the gel according to Schagger's Nature Protocol. I checked pH of every buffer (anode, cathode, gel buffer). My protein samples were dried well and resuspended in reducing buffer (the same as in the publication), so I think the influence of traces of alcohol can be omitted. Even ready-to-use protein marker floated out. Does anybody know what could cause such a problem?

Thank you in advance,

Kasia Roeske

do you flush the wells with electrode buffer immediately prior to loading the sample?

does your loading buffer contain glycerol or sucrose?

are you sure you diluted the electrode buffer to 1x?


my guess is that glycerol from the running gel is leaching into the wells so a flush prior to loading should clear the problem.

Check Loading Buffer again! Bromophenol Blue!

I saw this happen in our lab. The problem was that the researcher used 10X running buffer in the gel tank.