It depends on membrane pore size and protein Mw. If you use 0.45um membrane and your protein is small, then you might have protein escaping through the membrane. If in doubt, make sure to use 0.2 um membrane. Also, if you are afraid that proteins goes through the membrane, I guess transfer is complete in less than 2 hours, why not reducing transfer time.
Hi there,
2h transfer sounds long... I usually run wet transfer for 1h at 100V/ 350mA. Indeed proteins might pass beyond the membrane through its pores if running for too long. I would suggest you to shorten the transfer time.
Dear all,
In my 0.45 uM PVDF menbrane I see the loading control but I do not see any protein band after Ponceu staining. I transfer for 2 hours at RT at 200mV 2 gels at the same time. Do you think this might be too long?
Thank you in advance!
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