Handbook Of Biological Confocal Microscopy

pp 535-549

Multi-Photon Molecular Excitation in Laser-Scanning Microscopy

  • Winfried DenkAffiliated withMax-Planck Institute for Medical Research
  • , David W. PistonAffiliated withMolecular Physiology Biophysics, Vanderblt University
  • , Watt W. WebbAffiliated withSchool of Applied and Engineering Physics, Cornell University

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Multi-photon microscopy (MPM), which is based on molecular excitation by multi-photon absorption (MPA) and is usually combined with laser-scanning microscopy (LSM), has fulfilled its early promise (Denk et al., 1990), as evidenced by continued growth of its application to vital imaging of biological systems (for a recent collection of reprints, see Masters, 2003). Conventional fluorescence microscopy can provide submicron spatial resolution of chemical dynamics within living cells, but is frequently limited in sensitivity and spatial resolution by background due to out-offocus and scattered fluorescence.