In the surface plasmon resonance (SPR), a polarized light beam is reflected on the interface of a noble metal surface and a vicinal dielectric layer at an angle superior to the total internal reflection angle. Within this reflection process, a certain part of the light energy is coupled into an evanescent field that excites longitudinal charge density fluctuations of the free electron gas of the metal layer (surface plasmon waves) and propagates into the proximate dielectric medium with a decaying length of a few hundred nanometers. The incident angle of the light beam under which a maximum excitation of the surface plasmon waves occurs (surface plasmon resonance angle) is dependent on the wavelength of the exciting light source and the refractive index of the dielectric medium at the interface. Therefore, a variation of the optical properties and by this a change in refractive index of this dielectric layer will cause a measurable shift in the SPR angle or, if the incident angle is kept constant, a change in the intensity of the sensor reflected light.
A detailed overview of the SPR sensor technique and its bioanalytical applications is given in Robelek, R. (2009) Surface Plasmon Resonance Sensors in Cell Biology: Basics & Application. Bioanalytical Reviews, 1 (1), 57-72.
We use such a standard SPR sensor setup in combination with confluent layers of living, adherent, eukaryotic cells that can be cultured directly on top of the gold sensor surface. If the cells are stimulated and undergo biomolecular or morphologocal reactions, as it is for example the case in osmotic stress situations, a change in the refractive index of the cell elements residing within the evanescent field (cytosceleton, cytosol, membrane proteins etc.) can be followed in real time without the need of any disturbing label. Such sensors have for example been used by our group to perform label-free and time-resolved measurements of cell volume changes triggered by hypo- or hyperosmotic stimulations.
© 2008, Thomas Hirsch, Gisela Emmert.
