white bands in wb - (Dec/11/2006 )

hello all
i would appreciate ur help in this
in today's WB detection i found white bands in a very high background even in 1 seond exposure of the film?????
i didnt change any thing at all as yesterday for example....
i didnt change the first or second antibody or its concentartion or anything at all????
my samples are the same as before...
first time really to face this problem, i tried to wait 15 minutes after the first exposure and try again but my film was nothing there at all even the background.
i am using ECL detection .
i appreciate ur help.

This happened to me when once I switched from ECL to ECL advance (which is even more sensitive than ECL plus) without diluting enough my HRPsecondary. I got big white spots with less than 5 seconds exposure
So probably your white bands are also due to too much secondary, but if you didn´t change anything... it´s strange... Perhaps you took a wrong ECL (in case of somebody in the lab uses a more sensitive one and you confused with yours )
Let´s see what the others said.
In case it helps a bit, here it´s discussed this problem: http://www.protocol-online.org/biology-for...osts/11707.html

pumuki thankx
i know what u said and i did search the net for answers but the strange thing is that i didnt change anything from my usual and i was just repeating this western as i didnt include control in the last one ....
we are only 2 in the lab that use this ECL and its only one kit.
thankx for ur reply, waiting with u.

?

Could it be that your signal is now too high because you included your positive control whereas before you had your protocol titered to a lesser signal? Just guessing!