In this explorative study of the novel cationic lipid OO4 in two different formulations the complex formation with DNA, the biopharmaceutical stability of the lipid/DNA complexes in physiological media, and the transfection efficiency were analysed. We investigated liposomes composed of two binary mixtures of OO4 with either DOPE or DPPE as co-lipids in the molar ratio of 1:3. These formulations were compared with regard to their ability to bind the DNA using gel retardation electrophoresis, ethidium bromide exclusion and zeta potential measurements. Colloidal stability of the lipoplexes in foetal bovine serum (FBS) and the protective effect against degradation by endonucleases were studied. Furthermore, the influence of different salt concentrations on the complex formation with DNA was examined. The DOPE mixture was markedly superior compared to the DPPE mixture. Finally, haemocompatibility studies and gene silencing experiments were performed on OO4:DOPE 1:3 (n:n). The experiments demonstrate that the lipoplex formulation OO4:DOPE 1:3 (n:n) at N/P 4 is a promising candidate for systemic application because of the high colloidal stability in serum without PEGylated lipids, high transfection efficiency, superior resistance against nucleases, reproducible complexation independent of ionic effects, and haemocompatibility.