A series of peptide nucleic acid–oligo(bicycloguanidinium) (PNA–BG_n) conjugates were synthesized and characterized in terms of cellular antisense activity by using the pLuc750HeLa cell splice correction assay. PNA–BG₄ conjugates exhibited low micromolar antisense activity, and their cellular activity required the presence of a hydrophobic silyl terminal protecting group on the oligo(BG) ligand and a minimum of four guanidinium units. Surprisingly, a nonlinear dose–response with an activity threshold around 3–4 μm, indicative of large cooperativity, was observed. Supported by light scattering and electron microscopy analyses, we propose that the activity, and thus cellular delivery, of these lipo-PNA–BG₄ conjugates is dependent on self-assembled nanoaggregates. Finally, cellular activity was enhanced by the presence of serum. Therefore we conclude that the lipo-BG-PNA conjugates exhibit an unexpected mechanism for cell delivery and are of interest for further in vivo studies.