IR Surface Plasmon

Surface plasmon-based infrared spectroscopy for cell biosensing

Cell morphology is often used as a valuable indicator of the physical condition and general status of living cells. We demonstrate a noninvasive method for morphological characterization of adherent cells, see references. We measure infrared reflectivity spectrum at oblique angle from living cells cultured on thin Au film, and utilize the unique properties of the confined infrared waves (i.e., surface plasmon and guided modes) traveling inside the cell layer. The propagation of these waves strongly depends on cell morphology and connectivity. By tracking the resonant wavelength and attenuation of the surface plasmon and guided modes we measure the kinetics of various cellular processes such as (i) cell attachment and spreading on different substrata, (ii) modulation of the outer cell membrane with chlorpromazine, and (iii) formation of intercellular junctions associated with progressive cell polarization. Our method enables monitoring of submicron variations in cell layer morphology in real-time, and in the label-free manner.

Excitation of intracellular waveguide modes using a collimated broadband infrared beam. A| Experimental setup. A cell layer cultured on an Au-coated ZnS prism. The cells in the flow chamber exposed to culture medium at constant flow. The collimated and polarized infrared beam from the FTIR spectrometer impinges on the gold layer at angle θ_inc and excites surface plasmon and waveguide modes within the cell layer (panel C). The intensity of the reflected beam is measured by an MCT detector. The optical microscope placed on top follows the  viability of cell. B| Reflectivity spectrum from the ZnS/Au/MDCK cells/medium assembly at θ_inc=34.8⁰. The reflection minima correspond to the surface plasmon (SP) resonance and to the waveguide mode resonances (TM₁ and TM₂). C| Schematic representation of the electric field distribution for the surface plasmon and TM₁ waveguides modes. The surface plasmon penetrates only up to ~ 2 μm into the cell layer and is thus sensitive mainly to the cell-substrate interface. The TM₁ mode penetrates much further, it is confined within the entire cell volume and can be used to measure the cell height. D| Angular-resolved reflectivity spectra from the ZnS/Au/MDCK cells/medium assembly. The strong reflectivity minimum (deep blue) arises from the surface plasmon resonance. Its angular dependence mimics the dispersion of the water refractive index. A shallow minimum at lower angles (light blue) corresponds to the TM₁ waveguide mode. This mode does not appear in the absence of cell layer.

References
[1] Zilbershtein A, Golosovsky M, Lirtsman V, Aroeti B, et al. (2012) Quantitative surface plasmon spectroscopy: Determination of the infrared optical constants of living cells. Vibrational Spectroscopy 61: 43-49.
[2] Yashunsky V, Lirtsman V, Zilbershtein A, Bein A, et al. (2012) Surface plasmon-based infrared spectroscopy for cell biosensing. Journal of Biomedical Optics 17: 081409.[3] Yashunsky V, Lirtsman V, Golosovsky M, Davidov D, Aroeti B (2010) Real-time monitoring of epithelial cell-cell and cell-substrate interactions by infrared surface plasmon spectroscopy.Biophys J 99: 4028-4036.
[4] Yashunsky V, Shimron S, Lirtsman V, Weiss AM, Melamed-Book N, et al. (2009) Real-Time Monitoring of Transferrin-Induced Endocytic Vesicle Formation by Mid-Infrared Surface Plasmon Resonance. Biophysical Journal 97: 1003-1012.
[5] Golosovsky M, Lirtsman V, Yashunsky V, Davidov D, et al. (2009) Midinfrared surface-plasmon resonance: A novel biophysical tool for studying living cells. Journal of Applied Physics 105: 1020-1021.
[6] Ziblat R, Lirtsman V, Davidov D, Aroeti B (2006) Infrared surface plasmon resonance: a novel tool for real time sensing of variations in living cells. Biophys J 91:  776-776 .