FORMATION OF STABLE CATIONIC LIPID DNA COMPLEXES FOR GENE-TRANSFER/

Stable cationic lipid/DNA complexes were formed by solubilizing cation ic liposomes with 1% octylglucoside and complexing a DNA plasmid with the lipid in the presence of detergent. Removal of the detergent by di alysis yielded a lipid/DNA suspension that was able to transfect tissu e culture cells up to 90 days after formation with no loss in activity , Similar levels of gene transfer were obtained by mixing the cationic lipid in a liposome form with DNA just prior to cell addition. Howeve r, expression was completely lost 24 hr after mixing. The transfection efficiency of the stable complex in 15% fetal calf serum was 30% of t hat obtained in the absence of serum, whereas the transient complex wa s completely inactivated with 2% fetal calf serum. A 90-day stability study comparing various storage conditions showed that the stable comp lex could be stored frozen or as a suspension at 4 degrees C with no l oss in transfection efficiency. Centrifugation of the stable complex p roduced a pellet that contained approximately 90% of the DNA and 10% o f the lipid. Transfection of cells with the resuspended pellet and the supernatant showed that the majority of the transfection activity was in the pellet and all the toxicity was in the supernatant. Formation of a stable cationic lipid/DNA complex has produced a transfection veh icle that can be stored indefinitely, can be concentrated with no loss in transfection efficiency, and the toxicity levels can be greatly re duced when the active complex is isolated from the uncomplexed lipid.