Surface Plasmon Resonance Imaging for Cell Biology: Direct Measurement of the Evanescent Wave Penetration Depth

Abstract

Surface plasmon resonance imaging (SPRI) is a powerful label-free technique that has been known to provide sensitive biochemical surface measurements, but has only recently been applied to the field of cell biology. High resolution label-free imaging of cell-substrate contacts can be performed using a high numerical aperture objective lens. The SPRI signal is a result of the mass of material within the evanescent field, the refractive index of the material, and the distance between the material and the substrate. Cell focal contacts and cellular organelles can contribute to the SPRI signal. Unambiguous interpretation of the SPRI data is therefore complicated by uncertainties regarding the penetration depth of the plasmon wave. To gain a better theoretical understanding of the signal present in the image we used micron scale beads of polymers with a refractive index similar to cells as reference materials to calibrate SPRI reflectivity as a function of distance-to-substrate. Multi-wavelength measurements of these beads show that it is possible to tailor the effective depth of penetration of the evanescent wave into the cellular environment. The application of multiple wavelengths will assist the interpretation of the SPRI image.