My transfer buffer Ph is 8.7 but should be a 8.3 is this causing my exposures to be off also my bands look smeared. i am trying to troubleshoot and figure out why my chemiluminescence western blot are not coming out well. my antibodies are pyk2 and beta-tubulin.
Laura Pea
If using cell lysates as your sample I would suggest increasing the amount of lysis buffer - streaks can be an indication of incomplete digestion. You could also try letting the gel run at a lower voltage for a longer time to avoid streaking.
Also, have you tried increasing the amount of protein your are applying to your gel? 10 ng is a good start, but depending on the antibody I have gone up to 40 ng.
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