I have done a Southern Blot Analysis of DNAmt transferred to nylon membranes. The DNA was firstly loaded on a 2% agarose gel. An immunoassay was done to detect the bands using Anti-Digoxigenin antibody with Alkaline Phosphatase which catalyzes a colouring reaction with NBT and BCIP.
The results has a very strong background as you can see in the following pic:
I am not sure about what can be the reason of that background. I have read that thick agarose gels produce strong backgrounds, but I never experienced that before. If somebody has experience on this or alternative explanations, I would thank him/her to share them with me.