Radiant energy – Luminophor irradiation
Patent
1998-06-11
2000-12-26
Hannaher, Constantine
Radiant energy
Luminophor irradiation
2504591, 2504611, G01N 2139
Patent
active
061663859
DESCRIPTION:
BRIEF SUMMARY
BACKGROUND OF THE INVENTION
The present invention relates to a laser microscopy technique which produces molecular excitation in a target material by simultaneous absorption of three or more photons. The invention is an improvement over the two-photon laser microscopy technique disclosed in U.S. Pat. No. 5,034,613 to Denk et al. (hereinafter, the '613 patent), and this patent is hereby incorporated by reference.
The '613 patent discloses a laser scanning microscope which produces molecular excitation in a target material by simultaneous absorption of two photons to provide intrinsic three-dimensional resolution. Fluorophores having single photon absorption in the short (ultraviolet or visible) wavelength range are excited by a stream of strongly focused subpicosecond pulses of laser light of relatively long (red or infrared) wavelength range. The fluorophores absorb at about one half the laser wavelength to produce fluorescent images of living cells and other microscopic objects. The fluorescent emission from the fluorophores increases quadratically with the excitation intensity so that by focusing the laser light, fluorescence and photobleaching are confined to the vicinity of the focal plane. This feature provides depth of field resolution comparable to that produced by confocal laser scanning microscopes, and in addition reduces photobleaching. Scanning of the laser beam, by a laser scanning microscope, allows construction of images by collecting two-photon excited fluorescence from each point in the scanned object while still satisfying the requirement for very high excitation intensity obtained by focusing the laser beam and by pulse time compressing the beam. The focused pulses also provide three-dimensional spatially resolved photochemistry which is particularly useful in photolytic release of caged effector molecules.
A drawback to the two-photon laser microscopy technique disclosed in the '613 patent is that its applications are limited by the available laser technology. In particular, the two-photon technique requires use of a laser at specific wavelengths, depending upon the application, so that the sum of energy levels of the two photons provides the specific energy level needed to generate the desired fluorescent emission. Unfortunately, some laser microscopy applications would require use of a laser having a wavelength which is not technologically feasible at the present time. For example, excitation of chromophores that have very short wavelength absorption, such as amino acids and nucleic acids, would require a laser having a 540 nm wavelength using the two-photon technique, and such a laser does not exist at the present time.
SUMMARY OF INVENTION
The present invention provides a solution to the aforementioned problem through the application of three or more photon excitation to laser scanning fluorescence microscopy and to spatially resolved photo-chemical processing, such as caged reagent activation for micropharmacology and polymer cross linking for 3-d optical information storage.
Because three-photon induced fluorescence obeys a cubic dependence on excitation intensity and four photon excitation obeys a quartic dependence, both provide intrinsic three-dimensional resolution in laser scanning microscopy. Although such 3-d resolution has already been achieved by the nonlinear microscopy technique based on two-photon excitation disclosed in the '613 patent, three-photon excitation provides a unique opportunity to excite molecules normally excitable in the UV range (230-350 nm) with near IR light (700-1100 nm). Interesting biomolecules, such as the amino-acids tryptophan and tyrosine, the neurotransmitter serotonin and nucleic acids, have one-photon absorption peaks at approximately 260-280 nm, and fluorescence can be excited in these biomolecules by three and four photon excitation. The advantages of using long wavelength, near IR light are possibly less photodamage to living cells and conveniently available solid state femtosecond laser sources for deep UV absorbers. In practice, the
REFERENCES:
patent: 4170736 (1979-10-01), Wessel
patent: 4405237 (1983-09-01), Manuccia et al.
patent: 4407008 (1983-09-01), Schmidt et al.
patent: 4464761 (1984-08-01), Alfano et al.
patent: 4466080 (1984-08-01), Swainson et al.
patent: 4471470 (1984-09-01), Swainson et al.
patent: 4631581 (1986-12-01), Carlsson
patent: 4734578 (1988-03-01), Horikawa
patent: 4786170 (1988-11-01), Groebler
patent: 4791310 (1988-12-01), Honig et al.
patent: 4792341 (1988-12-01), Kozikowski et al.
patent: 4827125 (1989-05-01), Goldstein
patent: 4838679 (1989-06-01), Bille
patent: 4863226 (1989-09-01), Houpt et al.
patent: 4877965 (1989-10-01), Dandliker et al.
patent: 4887721 (1989-12-01), Martin et al.
patent: 5022757 (1991-06-01), Modell
patent: 5034613 (1991-07-01), Denk et al.
patent: 5196709 (1993-03-01), Berndt et al.
patent: 5239178 (1993-08-01), Derndinger et al.
patent: 5272089 (1993-12-01), Vo-Dinh
patent: 5289407 (1994-02-01), Strickler et al.
patent: 5376246 (1994-12-01), Page
patent: 5518694 (1996-05-01), Bentsen
patent: 5523573 (1996-06-01), Hanninen et al.
patent: 5777732 (1998-07-01), Hanninen et al.
Dimitri A. Parthenopoulos and Peter M. Rentzepis, "Three-dimensional Optical Storage Memory", Science, vol. 245 (Aug. 25, 1989), pp. 843-845.
Hollas 1987, "Modern Spectroscopy", John Wiley & Sons 1987, pp. 308-309.
Sheppard & Kompfner 1978, "Resonant scanning optical microscope", Applied Optic Col. 17 (Sep. 15, 1978), pp. 2879-2882.
Antonov et al. 1982, "Multiple Photon Processes in Molecules Induced by picosencond UV Laser Pulses", in Picosecond Phenomena III--Proceedings of the Third International Conference on Picosecond Phenomena, Jun. 16-18, 1982, pp. 310-314.
Gannaway & Sheepard 1978, "Second-harmonic imaging in the scanning optical microscope", Optical and Quantum Electronics vol. 10 (1978), pp. 435-439.
Wilson & Sheppard 1979, "Imaging and super-resolution in the harmonic microscope" Optica Acta, vol. 26 (1979), pp. 761-770.
Demptroder 1981, "Laser Spectroscopy", Springer-Verlag 1981, pp. 422-441 and 672-675.
Janis A. Valdmanis and R. L. Fork, "Design Considerations for a Photon Second Pulse Laser . . . ", IEEE Journal of Quantum Electronics, vol. QE 22, No. 1, Jan. 1986, pp. 112-118.
C.V. Shank "Generation of Ultrashort Optical Pulses", Topics in Applied Physics, vol. 60, Jan. 1988, pp. 5-31(particularly p. 15).
C.J.R. Sheppard: "Scanning optical microscope", Electronics and Power, pp. 166-172, Feb. 1980. Reprinted in SPIE Milestone Series, vol. MS 131, pp. 113-119, as enclosed.
Patrick P. Galmettes and Michael W. Berns: "Laser-induced multiphoton processes in living cells", Proc. Natl. Acad. Sci. USA, vol. 80, pp. 7197-7199, Dec. 1983, Biophysics.
He, et al., "Three-Photon-Absorption-Induced Fluorescence and Optical Limiting Effects in an Organic Compound," Optics Letters, vol. 20 (No. 14), p. 1524-26, (Jul. 15, 1995).
Davey, et al., "Three Photon Induced Fluorescence from a Conjugated Organic Polymer for Infrared Frequency Upconversion," Applied Physics Letters, vol. 67 (No. 7), p. 884-885, (Aug. 14, 1995).
Pantell, et al., "Theoretical and Experimental Values for Two-, Three-, and Four-Photon Absorptions," The Journal of Chemical Physics, vol. 46 (No. 9), p. 3507-3511, (May 1, 1966).
Wokosin, et al., "Multi-Photon Excitation Imaging with an All-Solid-State Laser," SPIE, vol. 2678, p. 38-49, (Jan. 31, 1996).
Ohsawa et al., "On the Possibility of Gas Temperature Measurement Using Two Photon Excitation," Proceedings of the 8th IMEKO Congress of the International Measurement Confederation, p. 523-528, (May 21, 1979).
Fritzler et al., "A Spectrometer for Semiautomatic Two Photon Fluorescence Spectroscopy," Journal of Physics E: Scientific Instruments, vol. 8 (No. 6), p. 530-532, (Jun. 30, 1975).
Slomba et al., "A Laser Flying Spot Scanner for Use in Automated Fluorescence Antibody Instrumentation," Journal of the Association for the Advancement of Medical Instrumentation, vol. 6 (No. 3), p. 230-234, (May 30, 1972).
Parthenopoulos et al., "Three Dimensional
Webb Watt W.
Xu Chris
Cornell Research Foundation Inc.
Hannaher Constantine
Israel Andrew
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