Despite the advantages of traditional fluorescence microscopy, the technique is hampered in ultrastructural investigations due to the resolution limit set by the diffraction of light, which restricts the amount of information that can be captured with standard objectives. In the past few years, a number of novel approaches have been employed to circumvent the diffraction limit, including near-field scanning optical microscopy (NSOM), stimulated emission depletion microscopy (STED), stochastic optical reconstruction microscopy (STORM) and structured illumination microscopy (SIM). These techniques have all achieved improved lateral (x-y) resolution down to tens of nanometers, more than an order of magnitude beneath that imposed by the diffraction limit, but each method has a unique set of limitations.
