RE:no bands after developing
Posted by: user (IP Hidden, Unregistered user, )
Date: July 27, 2004 08:18PM
>
> Hi
>
> I have used a stained gel[with Commassie] for western blot
> wet transfer.I am confident that bands transferred on to the
> membrane since I can visibly see them as the gel was
> stained.
>
> However, after Ab & secondary Ab treatments with washings &
> ECL kit ,I see absoultely nothing on my X-ray film.
>
> I have tried 15s,30s,1m,5m,15m exposures.
> I used HRPconjugated Ab's.
> Both primary & secondary Ab dilutions have been tried in 5%
> non fat dried milk in TBS-T & also in 0.05% milk in
> TBS-T.Blocking agent is 5% non fat dried milk.
>
> Any inputs on this one????
>
> Thank you
> Sari
>
hi sari,
i have had the same problem. maybe your AB are not specific enough. Also, see if your ECL kit is a little old.
 >  >
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