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Trouble when extracting Myosin Light Chain for WB - (Dec/07/2007 )

Hi everyone,

i'm attempting to run human T cells lysats on a gel and blot it against Myosin Light Chain (MLC2). Every time I've got important discrepancies of the MLC2 band intensities between wells. Yet, my loading control (ERK) is perfectly equal in all wells. Therefore I assume there's a problem with MLC2 solubilization in my lysis buffer (RIPA). Do you have an idea of what lysis buffer and protocols could I use to clear that hurdle?

thanks for your help


myosin requires high salt (0.3-0.6M) or atp (not sure of the concentration but mM range and still requires salt) to solubilize consistantly.