Removing endotoxin from plasmids

Mikhail Alexeyev malexe at biost1.thi.tmc.edu
Thu Aug 1 09:35:11 EST 1996


In article <msklar-3107962134520001 at msklar.cts.com>, msklar at cts.com (Mike
Sklar) wrote:

>      Hi.  I'm hoping someone can help me out.  I've been trying to purify
> endotoxin free plasmid DNA for in vivo work using a variety of methods. 
> I've tried using various kits and Phenol/chloroform to clean it up and
> nothing seems to work.  My  concentrations are still 2-5 EU/ug.  Can
> anyone offer a useful protocol or method for cleaning up the DNA?  I'd
> really appreciate it.
> 
> Thanks.
> 
> Mike Sklar

Qiagen sells special columns that allow to isolate DNA that is essentially
endotoxin-free. Double banding in CsCl also seems to work well, although
results are not very reproducible. You may also look at:

 Removal of endotoxins by affinity sorbents.   Anspach FB.  Hilbeck O.
      Journal of Chromatography A.  711(1):81-92, 1995 Sep 8.

  Bacterial lipopolysaccharide copurifies with plasmid DNA: implications
      for animal models and human gene therapy.   Wicks IP.  Howell ML.
      Human Gene Therapy.  6(3):317-23, 1995 Mar.

Lipopolysaccharide is a frequent contaminant of plasmid DNA
      preparations and can be toxic to primary human cells in the presence of
      Gene Therapy.  1(4):239-46, 1994 Jul.

Regards,
M.A.



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