6% gel westren blot not working as well as 10% - (Aug/07/2006 )
I am trying to western blot a protein that gets phosphoryl;ated. I identify the phosphorylation by bandshift. I use 6% gel to analyse this protein. But Imy 6% blotd look smeary and the samples are blurry and crooked. Thhe bandshifts get worse and worse. On the other hand, all higher % gels that I run like 10 or 12% run beautifully with sharp clear bands. can anyone help?
you shouldn't run the 6% at more than 80V for first hour. Then you can increase to 120 but not more...
and if your protein is in high MW, a 5V o/n semi dry transfert is very nice