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Principle of Western Bloting

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Dharvind Balan
Malaysia

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Topic Started by Dharvind Balan
on 7/31/2012 17:51 PM   
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 I have some question regarding the principle behind the protocols that we use for Western Bloting.

1) Why we need to soak PVDF membrane in methanol?
2) Why we need to arrange the blotting papers, gel and PVDF membrane in this order?
Blotting paper from anode 1, anode 2,PVDF, Gel, blotting paper from cathode buffer.
3) Purpose of blocking?
4) Adding solution 1 and 2 on top of PVDF membrane before placing a layer of plastic for film develop. What the solution will do?
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hcheung
Canada

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Posted By hcheung
on 8/1/2012 7:24 AM   
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Hi Dharvind Balan,

Sounds like you need to do some background reading. I would recommend:

1) "Western Blotting Handbook and Troubleshooting Guide"  from Pierce:

http://www.piercenet.com/Objects/View.cfm?Type=Page&ID=03971E57-D4D0-4B97-971C-0632E8224535

2) "Protein blotting handbook" from Millipore:

http://www.millipore.com/techpublications/tech1/tp001en00

3) "Western Blotting and Electrophoresis" from G Biosciences:

http://info.gbiosciences.com/complete-western-blot-handbook--selection-guide/
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