Dot blot- great, western blot - no signal - (Apr/22/2010 )

Hi.

I'm working on MMPs. When I did dot blots I recived great signal but when I tried when I tried western blots there was no signal at all. I don't know whats wrong cause I'm using the same conditions as in dot blots. I checked my transfer and it's ok.

What else might be wrong?

dot blots are usually of the native protein, western blots are of denatured protein. your antibody may not work with the denatured protein. if it is a commercial antibody, does the data sheet say that the antibody is certified for western blot?

Yes, antibodies are for western blot.

mdfenko on Apr 23 2010, 09:09 PM said:

the amount of protein in the band may be significantly lower than in the dot blot. you may need to incubate longer with and/or use a higher concentration of the antibody.