ECL Western troubles

Cara Baker crbaker at mail.tcd.ie
Fri Sep 6 03:01:19 EST 1996


Jose Bonner <jbonner at bio.indiana.edu> wrote:
>
> We're having trouble with Western blots probed by ECL.  Films sometimes 
> come up blank, even though antibodies and reagents are OK.  Sometimes a 
> 5-minute exposure is fine, but a 1-hour exposure (started just after) is 
> blank--not even any background.  What can interfere with ECL?  The 
> problem comes and goes; good blots interspersed with blanks.  Reprobe 
> the blanks, and sometimes they come up good; reprobe the good ones, 
> sometimes they come up blank.  Anyone else have this problem?
> 
> 

We have been using the ECL system with Western blots on nitrocellulose
and I have noticed that a tiny amount of fixer, even from a glove (fixer
contains ammonium thiosulphate, sodium acetate, boric acid and acetic
acid) will zap the ECL reagent, so fresh reagent has to be prepared in
a well-rinsed container.

Cara Baker
Dept. Biochemistry
TCD.



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