Surface-Sensitive Waveguide Imaging for In Situ Analysis of Membrane Protein Binding Kinetics

Ligand binding to membrane proteins initiates numerous therapeutic processes. Surface plasmon resonance (SPR), a popular method for analyzing molecular interactions, has emerged as a promising tool for in situ determination of membrane protein binding kinetics owing to its label-free detection, high surface sensitivity, and resistance to intracellular interference. However, the excitation of SPR relies on noble metal films, typically gold, which are biologically incompatible and can cause fluorescence quenching. Here we present a surface-sensitive waveguide imaging technique that retains the advantages of SPR and can be easily integrated into standard SPR devices by depositing additional dielectric layers onto SPR sensor chips using conventional vacuum evaporation. This novel sensor provides a silica surface, a well-known biologically compatible surface, for cell attachment and produces a sharper resonance curve and a stronger surface electric field than SPR, leading to improved measurement precision. After incorporating amplitude modulation, this approach increases measurement precision by approximately eight times compared to traditional SPR with the same optical setup. This high-precision method enables in situ single-cell analysis, revealing cell-to-cell heterogeneity that provides fuel for drug resistance. We believe this technique provides a streamlined strategy to enhance SPR for expanding their applications in biochemical research and drug screening.