Optics

Focus

Confocal theory

Dr. Marvin Minsky Confocal microscope: 1957 Confocal theory

LASER L ight A mplification by the S timulated E mission of R adiation Scanning Confocal Microscope Laser

Dichroic Mirror

Conventional / Confocal

Fluorescence 400 nm

Wavelength and Light Properties 575 nm E = h c   High Energy Low Energy 0,5* *2

Multiphoton exitation 800 nm 1200 nm 400 nm Dr. Maria Göppert-Mayer : theory of two-photon quantum transitions (two-photon absorption and emission) 1931,

LASER 500 nm Pulsed LASER 1000 nm1200 nm800 nm nm 500 nm Temporal photon concentration

Prof. Watt W. Webb et al. Two-photon laser scanning fluorescence microscopy : 1990 Spatial photon concentration

Confocal / Multiphoton

Luo at al. Cell Structure and Function 2006, 31: 63 Comparison of photoactivation of PA-GFP in vivo with single- photon (405 nm) and multiphoton (790 nm) laser light. Photoconversion Excitation area

Thorsten at al. Nature 2004; 427: 159 T-cell priming by dendriticcells in lymph nodes occurs in three distinct phases In VIVO Imaging