Phosphoinositide Thin layer chromatography question

Laura Saavedra via methods%40net.bio.net (by saavedralaura2004 from yahoo.es)
Mon Mar 5 16:39:14 EST 2007


Hi!

I am doing PIPK activity assay using PI4P lipid as substrate. I am doing the activity assays, extract the phsophoinositide lipids and analyze the formation of PI(4,5)P2 using TLC system with a solvent composition: CHCl3:CH3OH:NH4OH:H2O (90V/70V/4V/16V).
My question is that sometimes I have very nice sharp bands corresponding to PI(4,5)P2 and other times doing exactly the same procedure I obtained like a "wave" band of the lipid.
I would like to know what could be the problem? Is it any special requierement for prepearing the chamber? Or what could be the problem affecting the running? 

I would really appreciate any suggestion!

Thanks,

Laura
 		
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