Typically, endothelial cells are difficult to transfect. In this study, we report that antennapedia peptide (AP) linked to L-4F, a water-soluble, amphipathic a helical peptide that avidly binds lipids (AP-4F) increases Lipofectamine 2000-mediated transfection of bovine coronary endothelial cell cultures. Transfection efficiency was monitored by flow cytometry and fluorescent microscopy. Lipofectamine 2000 transfection of endothelial cell cultures with green fluorescence protein (GFP)-DNA typically yields transfection efficiencies of 35.4 +/- 3.3% with low levels of cell death (8.1 +/- 1.0%). Pre-treatment of the Lipofectamine 2000-GFP-DNA complexes with AP-4F for 5 min increased transfection to 58.2 +/- 2.8% without increasing cell death. AP-4F increases Lipofectamine 2000-mediated transfection in a time-dependent fashion (within 10-20 min). Systematic studies reveal that the individual components of AP-4F, i.e., AP and L-4F alone, are ineffective in increasing Lipofectamine 2000-mediated transfection and that AP-4F must be directly associated with DNA liposomes prior to transfection for optimal uptake by endothelial cells. These observations demonstrate that AP-4F may be useful for increasing the transfection efficiency of endothelial cell cultures with standard commercially available reagents. (C) 2003 Elsevier Science (USA). All rights reserved.