Fig. 4: Orientation of pre-adsorbed antibodies on the GA-lipo.

a Immunogold labeling showing the binding of pre-adsorbed TRA antibodies to secondary antibodies. b ζ potential of liposomes and TRA adsorption liposomes (n = 3 technical replicates, mean value ± s.d.). c Illustration of the orientation of pre-adsorbed antibodies on surface-galloylated liposomes. d Detection of TRA with fluorescently (FITC) labeled by flow cytometry. Data are shown as the amount of FITC-positive liposomes (n = 3 technical replicates, mean value ± s.d.). e Labeled adsorbed TRA with secondary-anti-human IgG1 targeting Fab of TRA measured by flow cytometry. Data are shown as the amount of PE-positive liposomes (n = 3 technical replicates, mean value ± s.d.). f AF488-labeled secondary-anti-human IgG1 targeting Fc of TRA. Data are shown as the amount of PE-positive liposomes (n = 3 technical replicates, mean value ± s.d.). Statistical significance (d–f) was determined by a two-way analysis of variance with Tukey’s multiple comparisons test. g Simulation snapshot of an equilibrated TRA protein bound to twenty GA-P1-Chol molecules (in purple). The ligand–protein interfaces are stabilized by hydrophobic pockets and hydrogen-bond networks, as indicated from domain 1 to domain 8, maintaining the structural integrity of TRA. h Time evolution of hydrogen bonds formed between GA-P1-Chol and the full-length trastuzumab (black) or its Fc region (red) during a 100 ns MD simulation. The increasing trend indicates progressive stabilization of the complex over time, with a higher number of hydrogen bonds observed in the full antibody structure. i Decomposition of binding energy contributions between GA-P1-Chol and each domain (1–8), showing Coulombic (electrostatic) and Lennard-Jones (van der Waals) interactions. j Time-resolved total interaction energy (kcal/mol) between GA-P1-Chol and individual antibody domains during the last 20 ns of MD simulation. Source data are provided as a Source Data file.