If the exciting radiation is halted, the fluorescence ceases. Interactive Java Tutorial Jablonski Diagram Explore how an electron absorbs energy, is excited into a higher energy state, and then decays. In some instances, the excited electrons may go from the triplet state to the lowest excited singlet state and then return to the ground state, emitting fluorescent light. Under some circumstances, e.g. photobleaching or the presence of salts of heavy metals, etc., emitted light may be significantly reduced or halted altogether, as discussed below. This reduction of emission intensity is generally called fading.
Some authors further subdivide fading into quenching and bleaching.
Bleaching is irreversible decomposition of the fluorescent molecules because of light intensity in the presence of molecular oxygen. Quenching also results in reduced fluorescence intensity and frequently comes about as a result of oxidizing agents or the presence of salts of heavy metals or halogen compounds. Sometimes quenching results from the transfer of energy to other so-called acceptor molecules physically close to the excited fluorophores, a phenomenon known as resonance energy transfer. This particular phenomenon has become the basis for a newer technique of measuring distances far below the lateral resolution of the light microscope. The occurrence of bleaching has led to a technique known as FRAP, or Fluorescence Recovery After Photobleaching .