Smear in WB - (Jun/05/2007 )

Dear all,

since a couple of days my western blots are a bit weird.
Basically everything is working, beside that my samples after the gel run and the wet blot look like smilies (bended downwards) as visible by Ponceau Red and after staining of course. This just happens recently and I never had this so far; so before going through all steps myself maybe anybody here had this annoying looking blots before and could give me an idea about what to fix?

Thanks a lot in advance
best regards!

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2 ideas:

running gels are becoming too hot during run; better cooling or less current may help;

irregular electric field: may be loss of running buffer at the upper edges of the gels

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[hi
i would use fresh running buffer with appropriate pH and run gel at low power.

good luck
sravan

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I had this happen not too long ago and I just made up new buffers for pouring the gels. I also purchased new TEMED because it tends to precipitate if old. Not sure if you pour your own gels but this seemed to work for me!

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It could also be a mismatch in the salt concentration of your sample vs the gel, or too much protein in the gel. Also, too much carbohydrates can cause it. These will cause problems that affect the shape of *each* lane, not *across* lanes. I'm not sure if your 'smile' shapes are across the whole gel, or individual 'smile' shaped bands.

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