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Staining of GLycoproteins in IEF


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#1 Prep!

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Posted 20 October 2009 - 11:52 PM

Hi All...
I have this confusion on the silver staining of glycoproteins in IEF gels. I have observed that with increasing glycosylation, the intensity of the glycoproteins after silver stain decreases. I attribute this to the masking of the aminoacids by the glycan chain resulting in the decrease in efficiency of the silver stain. Has anyone done an IEF for deglycosylated molecule? Does it give a more intense band than the glycosylated one? My next experiment would be that but if that is observed by most of you, I would not waste my resources on doing that. Kindly give some inputs..

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Pradeep
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#2 laurence.ylao

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Posted 26 October 2009 - 07:42 AM

Hi Pradeep,

I have no idea with IEF gel. But I tried the alcain Blue staining combined with silver staining together to see the glycoproteins. It works quite well.

Please see the paper as ref.

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Combined alcian blue and silver staining of subnanogram quantities of proteoglycans and glycosaminoglycans in sodium dodecyl sulfate-polyacrylamide gels.

Møller HJ, Heinegård D, Poulsen JH.

Anal Biochem. 1993 Feb 15;209(1):169-75.

PMID: 8465952
------------------------------------------------------------------

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Laurence

#3 Prep!

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Posted 26 October 2009 - 07:35 PM

hey thanx laurence.. will surely look into it..
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#4 heavensgate!

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Posted 03 November 2009 - 07:40 AM

hey thanx laurence.. will surely look into it..


Hi Pradeep,
I have worked with glycoproteins. They do stain differently compared non-glycosylated ones - they generally do not take up stain easily (may be for the same reason as you said). Generally antibody staining wroks better than silver, as it is more sensitive. You could try that as well.

#5 Prep!

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Posted 03 November 2009 - 07:19 PM

thanx heaven.. i will try that next.. but our lab still doesnt have a unit.. we are waiting for that only!!!
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