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Question about reducing agent (DTT) - (Jul/07/2010 )

Hi, i was preparing my protein samples earlier today and will load them on the gel tomorrow. While preparing my samples, i cannot recall if i have added the reducing agent (DTT) into my sample buffer used to lyse my cells with. I therefore plan to add DTT to my samples tomorrow (working concentration 50mM), heat them up and then load them on the gel.

My question is: If i have already added DTT previously and add DTT again tomorrow, will there be a negative impact on my results due to too much DTT? Could i 'over denature' my samples? :D


the samples will probably be able to handle the additional dtt but you should be able to smell if the samples already have dtt (especially at that high concentration).