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pakbiochemist

Member Since 16 Oct 2011
Offline Last Active Feb 24 2013 10:02 PM

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Active Posts 26
Profile Views 808
Member Title member
Age 29 years old
Birthday October 26, 1983
Gender
Female

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My research interests
biochemistry, proteomics, genomics

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Topics I've Started

membrane proteins ultrafiltration

24 February 2013 - 11:30 AM

Dear friends,

I have a very diluted sample of membrane proteins and i have to concentrate it. Can i use centricons of millipore or not as i have read that membrane proteins are very difficult to retain in their denatured state. is there any other method by which i can safely concentrate my protein sample.

Regards,

Binsan

suggestions for making use of tissue slides

18 October 2012 - 12:19 PM

Hi friends,

I am working on epigenetics(specifically Dna associated proteins). I have parrafinized tissue slides, please suggest me how can i make use of those slides in my work. simple histology has alraedy been done on those slides

Regards
Binsan

TAU gel electrophoresis of histones

01 October 2012 - 11:40 AM

Dear Friends,

I have run 2D of my histones (1D=Triton acid urea gel electrophoresis; 2D SDSPAGE). Please comment on the results in the attached file. I am not getting it.

Regards
Binsan

1.bmp 223.49K 77 downloads

Acid urea gel electrophoresis (AUGE)

04 September 2012 - 01:35 AM

Dear friends,

I am doing AU and TAU gel electrophoresis for some basic proteins by Sandra Hake Method "Extraction, purification and analysis of histones" Nature Protocols (2007). 2(6): 1445-1457.
Since the basic proteins are positively charged so they move in the opposite direction as that of SDS.

What i dont understand is that the author is asking to switch the electrode leads in Acid urea gel electrophoresis so that the positively charged proteins run towards the cathode. How should i switch the electrode leads? either the orientattion of leads going in the power supply or the electrode leads going in the tank, or there is anything else. I am using BioRad Mini Protean III apparatus. please help me i am really confused.

Regards

Binsan

Histone extraction from human tissue

02 August 2012 - 10:49 PM

Dear friends,

I have extracted histones from human tissues and have followed the following steps (Hake B, 2007)

1. PBS washing
2. Homogenization in lysis buffer (Tris, KCl, MgCl2, DTT, PMSF, Protease inhibitor, phosphatase inhibitor)
3. Centrifuge at 10,000g (10')
4. Pellet suspended in 0.4N H2SO4
5. Overnight incubation
6. Centrifugation at 16,000g (10')
7. Supernatant precipitated with TCA
8. Centrifuge at 16,000g (10')
9. pellet washed with acetone

My supervisor is saying that i cannot extract histones like this without first separating nuclei asmost of the histone extraction protocols uses sucrose gradientsfor nuclei isolation. Please guide me what i have extracted through this. Is this a crude extract or what?

Binsan
PhD Student

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