[3dem] Recalcitrant viruses

[Sacha De Carlo]

Hello all,
   
  whenever I have been working with polymerases (RNA pol II from Yeast or Human, or RNAP from Archaea) I always avoided glow-discharge, pre-soak the QF grids in chloroform for 30 min and let them dry in air prior to use. By following this very simple procedure I had RNA pols in the holes and not on carbon, on which they usually love to stick due to surface charges...
   
  Maybe it can help you...
   
  Best,
  Sacha
   
  

Eduardo Sanz-Garcia <esanzgar at chem.byu.edu> wrote:
  Hi all,

I am working with a virus that likes a lot the carbon and doesn't want 
to go to the holes.
Here are some pictures (the picture are not the best but it is possible 
to see the effect):
http://picasaweb.google.com/esanzgar/20080226?authkey=eoSXkRt-Ook

I have tried glow discharged and no glow discharged grids.
I have also tried different type of buffers, virus concentration, 
dialysis, and several applications of the virus in the grid.

Has anyone had a similar problem?
Does anyone have a solution for this problem?
Thank you very, very much

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==============================================
Sacha De Carlo, PhD
Molecular, Cell and Developmental Biology Department
Bio3D & Hoenger Labs, Campus Box 347
University of Colorado
Boulder, CO 80309

e-mail: sacha at bio3d.colorado.edu
URL: http://www.planetesacha.com
==============================================


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