SPR is a Quantum Mechanical Phenomenon
Snell’s law describes what happens when light is directed through a high refractive index prism (e.g. Sapphire - refractive index 1.76) to a surface of the prism in contact with a low refractive index medium, for example physiological buffer (See Figure 1). Light rays below the critical angle will exit the prism bending toward the prism surface. Light rays above the critical angle are totally internally reflected back through the prism. Photons that are totally internally reflected create an electric field at the interface. Light is not coming out of the prism but an electric field extends past the reflecting surface. This field oscillates with the usual characteristics of an electromagnetic mode. The electrical component perpendicular to the interface decays exponentially – this is called an evanescent wave. This wave is bound to the surface and is used in SPR to detect changes occurring at the surface.
SPR spectroscopy adds a thin metal layer between the prism surface and the aqueous compartment (See Figure 2). Free electrons in the metal layer can act as a resonator. Energy for the resonance comes from the evanescent wave produced by the totally internally reflected photons.
When certain conditions, determined by the wavelength, illumination angle, and refractive index of the prism, metal and aqueous layers are met, coupling/resonance occurs between the plasma oscillations of the free electrons in the metal and the bound electromagnetic field of the totally internally reflected photons. This coupling is the result of the momentum of the incoming light equaling the momentum of the plasma electromagnetic field. Photons are “absorbed” and converted to surface plasmons. Since the photons are not reflected, a “shadow” occurs in the reflected light.
Reichert Inc.’s SPR instruments are optically configured to illuminate a spot on the gold surface over a range of angles from about 58 to 85 degrees. When the aqueous compartment is e.g. physiological saline solution the “shadow” or SPR minimum occurs at about 66 Degrees. Mass changes at the interface between the gold layer and the aqueous compartment cause changes in the local refractive index near the gold layer. This changes the coupling/resonance angle. For example the if a protein layer was added to the gold / aqueous interface the coupling angle would be about 66.6 Degrees. Actual reflectivity from the SR7000 (Angle vs. reflectivity) before and after immobilizing a protein to the surface is shown in Figure 3.
In the SR7000 and SR7000DC Spectrometers the gold surface is illuminated with a range of angles that encompass these shifts in the coupling angle. This range of angles is continuously monitored with a linear photodiode array detector. Image analysis determines the angle at which the reflectivity minimum occurs. This is continuously sent to the data acquisition computer/software.
|
|
“The more success that quantum theory has, the sillier it looks”.
-Albert Einstein to Heinrich Zangger on Quantum Theory, May 20, 1912
“All these fifty years of conscious brooding have brought me no nearer to the answer to the question, 'What are light quanta?' Nowadays every Tom, Dick and Harry thinks he knows it, but he is mistaken. “
-Albert Einstein, 1954.
Figure 1
At incident angles below the critical angle light is refracted out of the prism, angle angles above the critical angle light is totally internally reflected back through the prism.
|
Figure 2
Adding a thin gold layer creates a “resonator” for the incoming light. This resonance occurs at an angle dependent on the refractive index or mass in contact with the solution side of the gold layer. As more mass is added to the surface, e.g. during an antigen binding to a surface immobilized antibody, the resonance angle “shadow” increases.
|
|
Figure 3
Reflected light image as viewed by the SR7000DC Photodiode Array image detector. The resonance angle increases as protein is added to the surface. This angle change is monitored and recorded in real-time. |
