We have developed a new method for observing cell/substrate contacts of living cells in culture based on the optical excitation of surface plasmons. Surface plasmons are quanta of an electromagnetic wave that travel along the interface between a metal and a dielectric layer. The evanescent field associated with this excitation decays exponentially perpendicular to the interface, on the order of some hundreds of nanometers. Cells were cultured on an aluminum-coated glass prism and illuminated from below with a laser beam. Because the cells interfere with the evanescent field, the intensity of the reflected light, which is projected onto a camera chip, correlates with the cell/substrate distance. Contacts between the cell membrane and the substrate can thus be visualized at high contrast with a vertical resolution in the nanometer range. The lateral resolution along the propagation direction of surface plasmons is given by their lateral momentum, whereas perpendicular to it, the resolution is determined by the optical diffraction limit. For quantitative analysis of cell/substrate distances, cells were imaged at various angles of incidence to obtain locally resolved resonance curves. By comparing our experimental data with theoretical surface plasmon curves we obtained a cell/substrate distance of 160 +/- 10 nm for most parts of the cells. Peripheral lamellipodia, in contrast, formed contacts with a cell substrate/distance of 25 +/- 10 nm.
Many plant pathogenic fungi, such as the cereal pathogen Colletotrichum graminicola , differentiate highly specialized infection structures called appressoria, which send a penetration peg into the underlying plant cell. Appressoria have been shown to generate enormous turgor pressure, but direct evidence for mechanical infection of plants by fungi is lacking. A microscopic method was developed that uses elastic optical waveguides to visualize and measure forces locally exerted by single appressoria. By this method, the force exerted by appressoria of C. graminicola was found to be about 17 micronewtons.
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