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Triplet Blinking stable triplet state S 1 T 1 due to solvent halogens transition metals

Transition to the Dark Side. Triplet Blinking stable triplet state S 1 T 1 due to solvent halogens transition metals. Quenching. Collision with solutes Resonance energy transfer. Energy transferred to non-fluorescent solute molecule. Fluorescence Resonance Energy Transfer.

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Triplet Blinking stable triplet state S 1 T 1 due to solvent halogens transition metals

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  1. Transition to the Dark Side • Triplet Blinking • stable triplet state • S1 T1 due to solvent • halogens • transition metals

  2. Quenching Collision with solutes Resonance energy transfer Energy transferred to non-fluorescent solute molecule

  3. Fluorescence Resonance Energy Transfer r < 50 A

  4. Death of a Fluor Photobleaching T1 is highly reactive O2 likes T1 electrons Oxygen radicals are highly reactive & cytotoxic O2 or other solute New compound (disrupted conjugation)

  5. Reflected Light (Epifluorescence) Microscopy

  6. Transmission Fluorescence Microscopy

  7. AOTF

  8. Widefield Fluorescence Microscopy

  9. Widefield Fluorescence Microscopy Confocal Fluorescence Microscopy

  10. Marvin Minsky’s Confocal Microscope Patent application submitted in 1957 while Minsky was a post-doc at Harvard

  11. Marvin Minsky’s Confocal Microscope Epi-illuminated design eliminated need for a condenser Also incorporated the concept of stage scanning

  12. ConfocalMicroscope Invented in 50s Not practical until availability of low cost lasers and high speed computers First commercial devices available in the 80s Uses pinhole to limit collected light to that which is “confocal” (i.e., shares conjugates planes of focus) with the object plane

  13. Zeiss LSM510

  14. Marvin Minsky’s Confocal Microscope Reduced blurring of image by light scattering Increased effective resolution Improved signal-to-noise ratio XY-scan possible across a wide area of the specimen Inclusion of Z-scan possibility Electronic adjustment of magnification Possibility of quantitative measurements of optical properties of the specimen

  15. Nipkow Disk Invented in 1884 by Paul Nipkow,a contemporary of Abbe. Central element of an early incarnation of the television.

  16. Tandem Scanning Optical Scopes

  17. Tandem Scanning Optical Scopes

  18. Light Amplification by Stimulated Emission of Radiation • Albert Einstein stimulated emission possible 1950s Charles Townes built first “MASER” Gordon Gould coined term LASER 1960 Theodore Maiman built first LASER (ruby)

  19. Stable LASER output requires establishment and maintenance of a “population inversion” of excitable electrons. Must “pump” the medium with either electrical or photonic energy

  20. ~ 10,000 - 40,000 hours

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