Staining nitrocellulose

*.* nikolaic at VISAR.WUSTL.EDU
Thu Mar 2 17:30:58 EST 1995


>Date:   26-FEB-1995 23:54
>From: Mary Jane Nather <natherm at ucs.orst.edu>
>Description: Staining nitrocellulose
>
>I am a microbiologist in a protein biochemistry lab, and I could really 
>use some help from anyone experienced in Western blotting, since no one 
>in this lab has experience working with antibodies.  
> 
>I have been looking for a procedure to stain nitrocellulose membranes 
>after transferring proteins from the polyacrylimide gel.  I have managed to 
>find out that I can use Coomassie, Amido Black, or Ponceau S, but I am 
>unable to determine the advantages to each method.  I don't necessarily 
>care if the stain is reversible, and we are amplifying the protein of 
>interest, so I don't think concentration should be a problem.
> 
>Any insights?  Does anyone have a preferred stain?
> 
>Thanks,
>MJ
> 
Hi, there. Below is what I gues coud be useful for you.

Notes:
i) The types of bond that hold proteins to nitrocel. are unknown. However,
the binding is blocked by oils or other proteins.
ii) Its recommended, the membrane to dry completely after transfer before
processing to blocking or staining. As alternative is to treat the membrane
with isopropanol for 1 min. If the dry membrane is processed wet it first
with water.

   Since, you don`t care whether or not stain is reversable, I would
recommend you to use Coomassie or Amido Black and forget about Ponceau.
Some people recommend Inda ink as superior over Amido Black staining but I
never used it.
Hope this helps.

Nikolai.

Nikolai
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If all fails, carefuly read the Manual
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