WB quick question - (Mar/28/2007 )
hello all
i have finished doing my western blot but found out that the developing machine is not working and need an hour from now to be fixed?
so can i still have good bands *i mean in density* after a whole one hour from ECL solution application to my membrane?
should i increase exposure time ? to what? 5 or 10 minutes?
thankxx
hi spanishflower!
that happened to me once and, unfortunately ECL (Amersham) signal dissapeared after 10-15 min 
even with long exposure times (10 min) I didn´t see anything.
Now I always check that the machine is switched on and works before applying ECL 
I have found this : http://www.protocol-online.org/forums/inde...mp;hl=ECL+again
hope this helps you
ohh
thankx
but i do always check and it was written that the maintinance is going to be from 9:30 am till 2:30 pm and its 3 pm here and the guy said he needs another hour
oh, that´s really annoying!
But you can try applying ECL again as someone suggested in the link I pasted.
pumuki
you will never belive it!!!
i went to try it anyway without adding ECL again and i exposue the film for 1 minute and 5 minutes and it was great !!!!
As if i did it on the spot!!!
thankx for your help really.
You´re welcome. You´re really lucky, congratulations!!! 
I´m glad for you.
Cheers 
sometimes re-applying ECL was not working (it was when I had weak bands and wanted to expose longer)
so, my suggestion for next time : expose a film few minutes as usually, and keep the film in a cassette protected from the light until you can reveal it. (I didn't try yet, but it should work)
hi missele how are you?
usually i do like what you said , i keep my membrane in my cassete closed all the time , may be this was the reason for yesterday's good results.
thankx again
usually i do like what you said , i keep my membrane in my cassete closed all the time , may be this was the reason for yesterday's good results.
thankx again
I'm fine, thank you.
I was busy, I had to finish things before to leave the lab for a few months.
I also keep the membrane in the cassette, but I was talking about the ECL film, because usually you expose just after adding the substrate and reveal immediately. but if the machine is not available, you can maybe wait between exposure and revelation