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3-dimensional confocal microscopy apparatus and focal plane scanning and aberration correction unit

Foreign code F130007601
File No. AF02-09WO
Posted date Aug 9, 2013
Country EPO
Application number 11824911
Gazette No. 2618197
Gazette No. 2618197
Date of filing Jul 27, 2011
Gazette Date Jul 24, 2013
Gazette Date Jun 8, 2016
International application number JP2011067719
International publication number WO2012035903
Date of international filing Jul 27, 2011
Date of international publication Mar 22, 2012
Priority data
  • P2010-208971 (Sep 17, 2010) JP
  • 2011JP067719 (Jul 27, 2011) WO
Title 3-dimensional confocal microscopy apparatus and focal plane scanning and aberration correction unit
Abstract Provided is a 3-dimensional confocal microscopy apparatus which is manufactured by combining a confocal microscope and an optical tweezers technique, wherein a pair of lenses for focal plane displacement where one lens is movable in the optical axis direction is arranged between a fixed objective lens and a fluorescent light imaging camera, and the 3-dimensional confocal microscopy apparatus also includes a mean which corrects the aberration of a fluorescent confocal image obtained by the fluorescent imaging camera.
Accordingly, it is possible to provide a 3-dimensional confocal microscopy apparatus which can acquire a 3-dimensional image of a specimen during a manipulation of the specimen using optical tweezers without affecting an optical trap.
Scope of claims [claim1]
1. A 3-dimensional confocal microscopy apparatus comprising: a light source for a first laser (1, 10) for confocal imaging excitation; a 2-dimensional scanning optical system for scanning the first laser on a specimen; an objective lens (2, 12) for focusing the first laser supplied from the light source on a fluorescent specimen which is an object to be observed; a fluorescent light imaging camera (16) for detecting a fluorescent confocal microscopy image based on a fluorescent light emitted from the fluorescent specimen; a pair of lenses (14, 15); lens drive means (17); a beam splitter or a dichroic mirror (20) and a light source (18) for a second laser; wherein
the pair of lenses (14, 15) for focal plane displacement is arranged between the objective lens and the fluorescent light imaging camera, the pair of lenses for focal plane displacement being constituted of an intermediate imaging lens (14) which forms an intermediate image of the fluorescent confocal microscopy image of the specimen and an intermediate objective lens (15) arranged such that a focal point of the intermediate objective lens overlaps with a focal point of the intermediate imaging lens on the same optical axis, and at least one lens out of these lenses being movable in an optical axis direction by the lens drive means (17),
the 3-dimensional confocal microscopy apparatus further includes a computer having an arithmetic operation means for acquiring a 3-dimensional structural image of the specimen by correcting aberration of the fluorescent confocal microscopy image caused by the movement of the lens of the pair of lenses for focal plane displacement in real time using the fluorescent confocal microscopy image acquired by the fluorescent light imaging camera and aberration correction data which is acquired in advance by performing provisional measurement of a sample specimen, and a means which displays a corrected image obtained by the arithmetic operation means, wherein
the beam splitter or a dichroic mirror (20) is provided between the pair of lenses for focal plane displacement and the objective lens, and the second laser supplied from the light source (18) for a second laser for optical tweezers passes through the objective lens via the beam splitter or the dichroic mirror and is focused on the specimen.
[claim2]
2. The 3-dimensional confocal microscopy apparatus according to claim 1, wherein the 2-dimensional scanning optical system includes a nipkow disc or a line scanning type scanning system.
[claim3]
3. The 3-dimensional confocal microscopy apparatus according to claim 1 or 2, wherein the lens drive means for the pair of lenses for focal plane displacement is one selected from a group consisting of a piezoelectric element, a magnetostrictive actuator and a voice coil.
[claim4]
4. The 3-dimensional confocal microscopy apparatus according to any one of claims 1 to 3, wherein timing for driving the lens drive means for the pair of lenses for focal plane displacement and timing for acquiring an image by a fluorescent imaging camera are adjusted so as to be synchronous with each other.
[claim5]
5. The 3-dimensional confocal microscopy apparatus according to any one of claims 1 to 4, wherein an arithmetic operation using an arithmetic operation means for acquiring a 3-dimensional structural image of a specimen is performed by a Graphics Processing Unit (GPU).
[claim6]
6. The 3-dimensional confocal microscopy apparatus according to any one of claims 1 to 5, wherein the 3-dimensional confocal microscopy apparatus further comprises a beam splitter or a dichroic mirror which is arranged between the objective lens and the pair of lenses for focal plane displacement, and an imaging lens and a bright field imaging camera for imaging a 2-dimensional image of a specimen separated by the beam splitter or the dichroic mirror.
[claim7]
7. A focal plane scanning and aberration correction unit to be incorporated into a 3-dimensional confocal microscopy apparatus comprises
lens drive means (17) and
a pair of lenses (14, 15) for focal plane displacement, the pair of lenses for focal plane displacement being constituted of an intermediate imaging lens (14) which forms an intermediate image of a fluorescent confocal microscopy image of a specimen and an intermediate objective lens (15) arranged such that a focal point of the intermediate objective lens overlaps with a focal point of the intermediate imaging lens on the same optical axis, and at least one lens out of these lenses being movable in an optical axis direction by the lens drive means, and
the focal plane scanning and aberration correction unit further comprises a computer having an arithmetic operation means for acquiring a 3-dimensional structural image of the specimen by correcting aberration of the fluorescent confocal microscopy image caused by the movement of the lens of the pair of lenses for focal plane displacement in real time using the fluorescent confocal microscopy image acquired by the fluorescent light imaging camera and aberration correction data which is acquired in advance by performing provisional measurement of a sample specimen, and a means which displays a corrected image obtained by the arithmetic operation means,
wherein the 3-dimensional confocal microscopy apparatus comprises a light source for a laser (10); an objective lens (12) for focusing the laser supplied from the light source on a fluorescent specimen; and a fluorescent light imaging camera (16) for detecting a fluorescent confocal microscopy image based on a fluorescent light emitted from the fluorescent specimen, and wherein
the focal plane scanning and aberration correction unit is to be arranged between the objective lens and the fluorescent light imaging camera.
  • Applicant
  • JAPAN SCIENCE AND TECHNOLOGY AGENCY
  • Inventor
  • IKUTA KOJI
  • IKEUCHI MASASHI
IPC(International Patent Classification)
Reference ( R and D project ) CREST Novel Measuring and Analytical Technology Contributions to the Elucidation and Application of Life Phenomena AREA
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