Nanoscopy with focused light

For more than a century, it has been generally accepted that the resolution of a lens-based optical microscope is limited to about d = λ /(2NA) > 200 nm in the focal plane and > 500 nm along the optic axis, with NA denoting the numerical aperture of the lens and λ the wavelength of light. The discovery in the 1990´s that elementary transitions between the states of a fluorophore can be used to eliminate the limiting role of diffraction has led to light microscopy concepts with resolution on the nanometer scale. Currently, all existing and successfully applied nanoscopy methods share a common enabling element: they switch fluorescence on or off, so that adjacent features are registered sequentially in time.