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1. (WO1991007651) TWO-PHOTON LASER SCANNING MICROSCOPY
Latest bibliographic data on file with the International Bureau

Pub. No.: WO/1991/007651 International Application No.: PCT/US1990/006482
Publication Date: 30.05.1991 International Filing Date: 13.11.1990
Chapter 2 Demand Filed: 01.05.1991
IPC:
G01N 21/64 (2006.01) ,G02B 21/00 (2006.01) ,G11B 7/004 (2006.01) ,G11B 7/12 (2012.01) ,G11B 7/135 (2012.01) ,G01N 21/63 (2006.01)
G PHYSICS
01
MEASURING; TESTING
N
INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
21
Investigating or analysing materials by the use of optical means, i.e. using infra-red, visible, or ultra-violet light
62
Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
63
optically excited
64
Fluorescence; Phosphorescence
G PHYSICS
02
OPTICS
B
OPTICAL ELEMENTS, SYSTEMS, OR APPARATUS
21
Microscopes
G PHYSICS
11
INFORMATION STORAGE
B
INFORMATION STORAGE BASED ON RELATIVE MOVEMENT BETWEEN RECORD CARRIER AND TRANSDUCER
7
Recording or reproducing by optical means, e.g. recording using a thermal beam of optical radiation, reproducing using an optical beam at lower power; Record carriers therefor
04
using fluorescent or phosphorescent record carriers
G PHYSICS
11
INFORMATION STORAGE
B
INFORMATION STORAGE BASED ON RELATIVE MOVEMENT BETWEEN RECORD CARRIER AND TRANSDUCER
7
Recording or reproducing by optical means, e.g. recording using a thermal beam of optical radiation, reproducing using an optical beam at lower power; Record carriers therefor
12
Heads, e.g. forming of the optical beam spot or modulation of the optical beam
G PHYSICS
11
INFORMATION STORAGE
B
INFORMATION STORAGE BASED ON RELATIVE MOVEMENT BETWEEN RECORD CARRIER AND TRANSDUCER
7
Recording or reproducing by optical means, e.g. recording using a thermal beam of optical radiation, reproducing using an optical beam at lower power; Record carriers therefor
12
Heads, e.g. forming of the optical beam spot or modulation of the optical beam
135
Means for guiding the beam from the source to the record carrier or from the record carrier to the detector
G PHYSICS
01
MEASURING; TESTING
N
INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
21
Investigating or analysing materials by the use of optical means, i.e. using infra-red, visible, or ultra-violet light
62
Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
63
optically excited
Applicants:
CORNELL RESEARCH FOUNDATION, INC. [US/US]; Suite 105 20 Thornwood Drive Ithaca, NY 14850, US
Inventors:
DENK, Winfried; CH
STRICKLER, James, P.; US
WEBB, Watt, W.; US
Agent:
COOPER, George, M. ; Jones, Tullar & Cooper 2001 Jefferson Davis Highway Suite 1002 Arlington, VA 22202, US
Priority Data:
436,04514.11.1989US
Title (EN) TWO-PHOTON LASER SCANNING MICROSCOPY
(FR) MICROSCOPIE A BALAYAGE LASER A DEUX PHOTONS
Abstract:
(EN) A laser scanning microscope (10) produces molecular excitation in a target material by simultaneous absorption of two photons to thereby 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 (14) 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 strongly focusing the laser light (14), fluorescence as well as photobleaching are confined to the vicinity of the focal plane (18). 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 (14), by a laser scanning microscope (10), 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 (14) 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.
(FR) Un microscope à balayage laser (10) provoque l'excitation moléculaire d'un matériau cible par absorption simultanée de deux photons entraînant ainsi une résolution tridimensionnelle intrinsèque. Des fluorogènes présentant une absorption de photons unique dans les longueurs d'ondes courtes (ultraviolet ou visible) sont excités par un flux d'impulsions (14), inférieures à la picoseconde et fortement focalisées, de lumière laser d'étendue de longueur d'onde relativement longue (rouge ou infrarouge). Les fluorogènes absorbent à au moins une demie longueur d'onde laser de manière à produire des images fluorescentes de cellules vivantes et d'autres objets microscopiques. L'émission fluorescente provenant des fluorogènes augmente de façon quadratique avec l'intensité d'excitation de telle sorte qu'en focalisant fortement la lumière laser (14), la fluorescence aussi bien que le photoblanchissement sont limités au niveau du plan focal (18). Cette caractéristique fournit une profondeur de résolution de champ comparable à celle produite par des microscopes à balayage laser à foyer commun, et de plus réduit le photoblanchissement. Le balayage du faisceau laser (14), par un microscope à balayage laser (10), permet une construction d'images par réception de fluorescence causée par deux photons à partir de chaque point dans l'objet balayé tout en satisfaisant l'exigence d'une intensité d'excitation très élevée obtenue par la focalisation du faisceau laser (14) et par la compression d'impulsions du faisceau. Les impulsions focalisées assurent également une photochimie à résolution spatiale tridimensionnelle qui est particulièrement utile dans la libération photolytique de molécules effectrices en cage.
Designated States: JP
European Patent Office (AT, BE, CH, DE, DK, ES, FR, GB, GR, IT, LU, NL, SE)
Publication Language: English (EN)
Filing Language: English (EN)
Also published as:
EP0500717