[Protein-analysis] NOS western blot problem

Wang, Hongyan via proteins%40net.bio.net (by Hongyan.Wang At umassmed.edu)
Sat Feb 3 20:43:44 EST 2007


Hi, Anna,

 

I found that I now have the same problem as you had, except the gene
that do well is SOD1, actin as housekeeping gene.  Could you please tell
me what happened to your samples and how did you solve it.  I would be
grateful for suggestions. Thanks.

 

Hongyan

 

Anna Durrans anna.durrans at physiol.ox.ac.uk
<mailto:proteins%40iubio.bio.indiana.edu?Subject=NOS%20western%20blot%20
problem&In-Reply-To=> 
Fri Jul 13 08:39:40 EST 2001 

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________________________________

Hi everyone,
 
I have been running westerns for eNOS, nNOS and iNOS successfully using
various rodent organ tissues. I use actin as a housekeeper.
 
In order to show reproducibility I recently re-ran the exact same
lysates
again, along with some freshly made lysates. Strangely I found that the
actin signal was very much weaker in some (not all) of the original
lysates,
as wells as in the fresh lysates.
 
I re-ran all these again, and while there is a pretty stong actin
signal,
it
is still weaker in those tissues which before showed a weak actin
signal.
 
I am not sure what could have happened. The NOS signal in ALL the
tissues
was fine - so it is not the lysates themselves but possibly the actin
specifically which has degraded.
 
Has anyone else had this problem? Does anyone know what could have
happened?
 
I would be really grateful for any suggestions.
 
Anna
 
 
 
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