Hi there
I have a Myc tagged protein. When I first received the plasmid, transfected, etc, I was able to see the Myc tagged protein band during Western, but now when I run it, I am not able to see the band. When I blot with the antibody raised against my specific protein, I am able to pick up the band with no issue.
Is there any reason why the myc Ab won't detect my myc tagged protein?
I thought I was diluting it too much but even with a small amount, I still can't get anything.
Any comment's would be greatly appreciated!
Thank you
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3 replies to this topic
#2
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Hmmm, I think it's working
Posted 11 April 2011 - 03:47 PM
Antibody has gone off? Do you have another protein that is also myc tagged that you can use to check whether it is the antibody or the protein?
#3
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Enthusiast
Posted 12 April 2011 - 05:07 AM
There is a possibility that during this time I have been working with my protein, the myc tag might have broken off?
#4
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Hmmm, I think it's working
Posted 12 April 2011 - 03:32 PM
Not necessarily broken, but perhaps your plasmid is degraded by freeze/thaw or DNAse action. It could also be that the plasmid has been mutated by the bacteria while you were growing it up.
