TIRF operates on the principle of total internal reflection. When a light beam passes from a medium of higher refractive index to one of a lower refractive index at an angle greater than the critical angle, it undergoes total internal reflection. An evanescent wave is generated at the interface, which decays exponentially with distance from the surface. This evanescent wave can excite fluorophores close to the interface, allowing for high-resolution imaging of surface phenomena.
