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doublets on 2D gel


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#1 biochem06

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Posted 09 March 2016 - 06:57 AM

Hi,

 

What would cause the "doublets" that I'm seeing on my 2D gels?

 

thanks

 

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#2 mdfenko

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Posted 09 March 2016 - 12:55 PM

i would think that would be proteins with similar pI and a little different molecular weight.

 

you can check them on mass spec.

 


Edited by mdfenko, 09 March 2016 - 12:56 PM.

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#3 biochem06

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Posted 14 March 2016 - 01:30 AM

I remember reading somewhere (but can't find it now) that it can be an artifact. Due to placement of the strip on the gel?



#4 biochem06

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Posted 14 March 2016 - 01:42 AM

Ah, I just found something, although it's still unclear why it happens.

From - http://www.bio-rad.c...lletin_2651.pdf

3. Twin vertical spots or vertical doublets throughout the gel.
This effect is sometimes seen when thiourea is included in the sample solution. Its cause is unclear.


#5 Michael Starr

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Posted 28 March 2016 - 02:29 PM

My GUESS (from very limited background of OChem) is that the sulfur in the thiourea reacts with a population of the protein, on the cysteine, to form a "post-translational modification" disulfide bond that modifies the molecular weight of the protein.

 

What is the concentration of thiourea in your sample? Any chance of calculating the molar ratio of thiourea/protein to see if we're using all the thiourea up and have excess protein?

 

Admins feel free to tell me I'm crazy.


Edited by Michael Starr, 28 March 2016 - 02:30 PM.





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