High Resolution Surface Plasmon Microscopy: From Nano-Colloids To Single Nucleosome Imaging

Surface plasmon resonance (SPR) has been widely recognized as a highly sensitive and non intrusive method for probing modifications of adsorbed layers, and in particular for its application to characterize biomolecular specific interactions such as antigen-antibody recognition. We apply SPR to study nucleosomes, first level of DNA compaction around an octamer of proteins called histones. To maintain the DNA helix accessible to the transcription and replication machineries during the cell cycle, this complex is highly dynamical (formation, disassembly, or sliding of the nucleosome), leading to rapid modifications of the whole chromatin structure in vivo. We aim at understanding how the DNA sequence influences the structure and dynamics of the nucleosomes in chromosomes. The scanning surface plasmon microscope (SSPM) set-up relies on the use of a high numerical aperture objective that confines the surface plasmon polaritons (SPPs) to an area of the interface much smaller (up to a few hundreds of nanometers) than their typical propagation length (few microns). Similarly to the Kretschmann configuration, SPPs are excited at the resonance angle θP. As an objective is used to focus the purely P-polarized (radial) light, the SPPs converge to the center of the illuminated area, leading to the creation of SPPs interferences that will reradiate in symmetrical rays and go through the objective to be detected. The presence of non-marked biological samples at the interface modifies the propagation conditions (i.e. the interferences) of the SPPs, leading to a change in the image contrast. The SSPM point spread function is about 150 nm in aqueous medium, providing high resolution images of biological samples in vitro. We will present the SSPM study of the optical response of gold and latex nanoparticles and then the first images of non-marked single nucleosomes in liquid medium.