Antisense oligo experiments - help

Wolfgang Schechinger u7k0201 at sunmail.lrz-muenchen.de
Mon Jul 17 12:42:47 EST 1995


grggta at picr.cr.man.ac.uk (Graham Atherton) wrote:
>
>Has anyone experience of using 22mer antisense oligo's to give 
>biological effects when added to adherent mammalian cells eg interfere
>with cell cycle. All my experiments so far show as much effect when
>sense (control) oligo's are used as when antisense are used.
>I just add phosphorothioate oligo's to the growth medium at 5-15uM.
>Would it be better to use lipofectin (or similar) to aid uptake of the
>oligo and hence increase specific effect - does anyone have a good
>protocol for this? 
>
>Cheers
>Graham
>
Dear Graham, 

I`m doing antisense experiments in a similar way. I`m using fully thioated 17mers against a 
G-Protein added to the culture medium in concentration from 0,2 to 2 µM. I'm testing time- and 
concentration dependencies and am using several controls (sense, modified, mixed and scrambled 
sequences)


In our institue, we made best experience with the last trity protection group left on the 
molecule for getting it better into the cells. A colleague made experiments with lipofectin 
too, but he could not see any enhancement of uptake. Some authors packed the oligos into 
liposomes and achived good results. On cells owning specific receptors (e.g. folate on liver 
cells) you even could use liposomes carrying an 'antenna molecule' for cell specific directing 
and enhancing uptake by selective endocytosis of the loaded lipose.

Maybe this helps

++++++++++++++++++++++++++++

Wolfgang Schechinger
Instite for Diabetes Reserch
Kölner Platz 1
80804 Munich
Phone +49 (89) 30 79 31 24
Fax   +49 (89) 30 81 733
email u7k0201 at sunmail.lrz-muenchen.de




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