Monitoring Molecular Interactions with Cell Membranes Using Time-Dependent Second Harmonic Generation Microscopy

Time-resolved second harmonic generation (SHG) microscopy is used to investigate the physicochemical interactions between positively charged, hydrophobic, drug-like molecules and the plasma membrane of human cells (nonsmall cell lung cancer, H596). In the present study, molecular adsorption and transport of the cationic molecules, malachite green (MG) and malachite green isothiocyanate (MGITC), are studied in real time in living H596 cells and in dead, fixed H596 cells. MGITC is shown to have stronger adsorption and more rapid transport kinetics as compared to MG due to increased dipole–dipole interactions. Additionally, MGITC is found to have faster adsorption and transport kinetics in living H596 cells in comparison to fixed H596 cells, as well as higher dispersity in transport rate, pointing to changes in the nature of the plasma membrane or its integrity. Overall, the findings highlight the importance of electrostatic interactions, chemical functional groups, and cell integrity in molecular translocation dynamics across cell membranes.