western blot problems
Posted 21 April 2009 - 09:21 PM
Posted 22 April 2009 - 12:25 PM
For faint bands, you could try exposing the film longer, but I suspect you may just have a bad Ab. Has anyone in your lab used this Ab for Westerns before? The white blotches are likely a result of pipeting the detection agent directly onto the surface of the membrane. Try pipeting it onto the edge of the membrane, then tipping it to disperse over the whole membrane.
then heaven will be yours, before you meet your end
Posted 22 April 2009 - 05:00 PM
It seems kind of obvious, but how big is your protein? Are you transferring for long enough to get it onto your membrane?
From your description it does sound like you have a little problem with transfer, partial bands are usually the result of bubbles (maybe in the pads, not between the gel and membrane).
Posted 23 April 2009 - 06:54 AM
I would also guess that this is a case of "bad Ab". Is there any way you can try a positive control blot, such as actin?