How the West(ern) was lost - (Aug/29/2001 )

Could someone suggest why, after developing a blot using PVDF membrane, polyclonal antibodies, HRP conjugate and 4C1N substrate, the resulting bands appear a muddy brown and fade rapidly into oblivion? Previous ELISA experiments have demonstrated a stong affinitiy for the antigen by the antibodies.

I would appreciate any wisdom.

Nick

try changing your peroxide or the substrate. I had been through those misterious days until i changed over to Amersham ECL developing kit.After secondary-HRP wash your blot and develop with the mixture and expose to x-ray sheet. Will get good results. DOnt have to worry where your bands dissappears

try changing your peroxide or the substrate. I had been through those misterious days until i changed over to Amersham ECL developing kit.After secondary-HRP wash your blot and develop with the mixture and expose to x-ray sheet. Will get good results. DOnt have to worry where your bands dissappears

You may also want to try Nitrocellulose membrane. I always had better results with Nitrocellulose than PVDF membranes. Biorad sells a good Nitro- membrane.