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2 bands very close in the expected place

WB native denaturating

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#1 madelingirly

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Posted 15 March 2016 - 10:09 PM

Dear all,

I have done WB, but I did denaturing of my sample for only 3 minutes (I used sds-RIPA as lysis buffer).

So can these very close 2 bands be the native and denaturing protein, since I did not do the whole recommend time (5 minutes).

Thanks for any suggestion

Mady

 

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#2 GNANA

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Posted 16 March 2016 - 06:00 AM

is this a receptor or any membrane protein? have you ever done this and had only single band? Looking at the band pattern to me the two bands are a post-translational modified form of the same protein most likely Fully glycosyated vs non/partial glycosylated protein, or phosphorylation and non-phosphorylated. ofcourse you can confirm by denaturing at a longer time.


I always had an alternate hypothesis....


#3 madelingirly

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Posted 18 March 2016 - 12:18 AM

Thanks GNANA for ur comment, this is true a membrane protein.

anyway, I tried to increase the denaturation time as recommended, then I would update here .

Mady







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