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Differentiating between long and short isoforms of Dopamine D2 Receptor using We

D2R Rat Striatum Western Blotting

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#1 Heli

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Posted 19 September 2013 - 10:12 AM

Hello,

We are trying to assess levels of D2 Receptor short form in rat striatal synaptosomes using Western blotting. However there are no specific antibodies to differentiate between the two. The bands we get are too smeared too because of the different glycosylated forms of both the isoforms. Has someone dealt with D2R isoforms before? Any help with suggesting antibodies or a different fractionation protocol or any other technique would be appreciated...Thanks

Heli



#2 jerryshelly1

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Posted 19 September 2013 - 10:58 AM

You could enzymatically deglycosylate your proteins.  It wouldn't be 100%, but you should see an relative increase in your band intensities. You could convince your reviewers on the effectiveness with any RT-PCR data you have on the isoforms.

 

NEB enzyme people have used in the lab (I'm sure others exist).

https://www.neb.com/...ycosylation-mix

 

Good luck

 

Edit-You are probably already aware, but you may be able to get a free sample to convince your PI it works before purchasing.


Edited by jerryshelly1, 19 September 2013 - 10:59 AM.






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