Separating proteing with difference of 5Kd on western blot

1D, 2D, HPLC, SDS PAGE, etc. Talk about it here.
Preeti
Carbon Member
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Joined: Mon Dec 17, 2012 8:08 pm

Separating proteing with difference of 5Kd on western blot

Postby Preeti » Mon Dec 17, 2012 8:14 pm

Hello everyone,
I am kind of stuck here with two proteins with almost similar molecular weight and I want to do Western blot of them as it is a
requirement of the project. Can someone guide me?? the weight is 90kd and 95 kd.
Thanks,
Preeti

Biomarker
Albumin Member
Posts: 88
Joined: Thu Aug 09, 2012 3:28 am

Hi

Postby Biomarker » Tue Dec 18, 2012 12:02 am

Hi Preeti,

What percentage of SDS-PAGE you are using? Try to alter the percentage of acrylamide. I hope this will help to you..

Biomarker

Preeti wrote:Hello everyone,
I am kind of stuck here with two proteins with almost similar molecular weight and I want to do Western blot of them as it is a
requirement of the project. Can someone guide me?? the weight is 90kd and 95 kd.
Thanks,
Preeti

Preeti
Carbon Member
Carbon Member
Posts: 4
Joined: Mon Dec 17, 2012 8:08 pm

Postby Preeti » Tue Dec 18, 2012 3:03 pm

Hello Biomarker,
I am using 4-17% gradient gel as i have four more proteins which I need to study on same blot. All to gather i am studying six proteins and one as control common housekeeping protein.
If you have any idea to sort this problem please do snuggest me. Thanks for your reply.
:) Preeti

Sidney
Glycine Member
Glycine Member
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Joined: Sun Dec 04, 2011 4:41 am

Postby Sidney » Tue Dec 18, 2012 6:50 pm

Hi, Preeti,

I would like to suggest you perform western blotting from two SDS-pages, because antibody has very high specificity to assure the right protein being annalyzed.

Sid

Biomarker
Albumin Member
Posts: 88
Joined: Thu Aug 09, 2012 3:28 am

Hi

Postby Biomarker » Wed Dec 19, 2012 2:23 am

Hi Preeti,

Try 8-16% and 10-16%.
Good luck....

Cheers...
Biomarker

Preeti
Carbon Member
Carbon Member
Posts: 4
Joined: Mon Dec 17, 2012 8:08 pm

Postby Preeti » Wed Dec 19, 2012 4:24 pm

Thanks for your suggestion Biomarker!!! :)
Would inform you the outcome and I wish either of them will work!!!
Cheers, :D

Preeti

Preeti
Carbon Member
Carbon Member
Posts: 4
Joined: Mon Dec 17, 2012 8:08 pm

Postby Preeti » Wed Dec 19, 2012 5:50 pm

Thanks for your suggestion Biomarker!!! :)
Would inform you the outcome and I wish either of them will work!!!
Cheers, :D

Preeti


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