WO2014131853A1 - Microscope doté d'un objectif optique et acoustique combiné - Google Patents

Microscope doté d'un objectif optique et acoustique combiné Download PDF

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Publication number
WO2014131853A1
WO2014131853A1 PCT/EP2014/053875 EP2014053875W WO2014131853A1 WO 2014131853 A1 WO2014131853 A1 WO 2014131853A1 EP 2014053875 W EP2014053875 W EP 2014053875W WO 2014131853 A1 WO2014131853 A1 WO 2014131853A1
Authority
WO
WIPO (PCT)
Prior art keywords
sample
pressure sensor
microscope according
module
objective
Prior art date
Application number
PCT/EP2014/053875
Other languages
German (de)
English (en)
Inventor
Tobias Schmitt-Manderbach
Wibke Hellmich
Gerhard Krampert
Helmut Lippert
Original Assignee
Carl Zeiss Ag
Carl Zeiss Microscopy Gmbh
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Carl Zeiss Ag, Carl Zeiss Microscopy Gmbh filed Critical Carl Zeiss Ag
Publication of WO2014131853A1 publication Critical patent/WO2014131853A1/fr

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Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N29/00—Investigating or analysing materials by the use of ultrasonic, sonic or infrasonic waves; Visualisation of the interior of objects by transmitting ultrasonic or sonic waves through the object
    • G01N29/04—Analysing solids
    • G01N29/06—Visualisation of the interior, e.g. acoustic microscopy
    • G01N29/0654—Imaging
    • G01N29/0681—Imaging by acoustic microscopy, e.g. scanning acoustic microscopy
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N29/00—Investigating or analysing materials by the use of ultrasonic, sonic or infrasonic waves; Visualisation of the interior of objects by transmitting ultrasonic or sonic waves through the object
    • G01N29/04—Analysing solids
    • G01N29/06—Visualisation of the interior, e.g. acoustic microscopy
    • G01N29/0654—Imaging
    • G01N29/069—Defect imaging, localisation and sizing using, e.g. time of flight diffraction [TOFD], synthetic aperture focusing technique [SAFT], Amplituden-Laufzeit-Ortskurven [ALOK] technique
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N29/00—Investigating or analysing materials by the use of ultrasonic, sonic or infrasonic waves; Visualisation of the interior of objects by transmitting ultrasonic or sonic waves through the object
    • G01N29/22—Details, e.g. general constructional or apparatus details
    • G01N29/24—Probes
    • G01N29/2418—Probes using optoacoustic interaction with the material, e.g. laser radiation, photoacoustics
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/0004—Microscopes specially adapted for specific applications
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/02—Objectives
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/24—Base structure
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01S—RADIO DIRECTION-FINDING; RADIO NAVIGATION; DETERMINING DISTANCE OR VELOCITY BY USE OF RADIO WAVES; LOCATING OR PRESENCE-DETECTING BY USE OF THE REFLECTION OR RERADIATION OF RADIO WAVES; ANALOGOUS ARRANGEMENTS USING OTHER WAVES
    • G01S15/00—Systems using the reflection or reradiation of acoustic waves, e.g. sonar systems
    • G01S15/88—Sonar systems specially adapted for specific applications
    • G01S15/89—Sonar systems specially adapted for specific applications for mapping or imaging
    • G01S15/8906—Short-range imaging systems; Acoustic microscope systems using pulse-echo techniques
    • G01S15/8909—Short-range imaging systems; Acoustic microscope systems using pulse-echo techniques using a static transducer configuration
    • G01S15/8915—Short-range imaging systems; Acoustic microscope systems using pulse-echo techniques using a static transducer configuration using a transducer array
    • G01S15/8922—Short-range imaging systems; Acoustic microscope systems using pulse-echo techniques using a static transducer configuration using a transducer array the array being concentric or annular
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01S—RADIO DIRECTION-FINDING; RADIO NAVIGATION; DETERMINING DISTANCE OR VELOCITY BY USE OF RADIO WAVES; LOCATING OR PRESENCE-DETECTING BY USE OF THE REFLECTION OR RERADIATION OF RADIO WAVES; ANALOGOUS ARRANGEMENTS USING OTHER WAVES
    • G01S15/00—Systems using the reflection or reradiation of acoustic waves, e.g. sonar systems
    • G01S15/88—Sonar systems specially adapted for specific applications
    • G01S15/89—Sonar systems specially adapted for specific applications for mapping or imaging
    • G01S15/8906—Short-range imaging systems; Acoustic microscope systems using pulse-echo techniques
    • G01S15/899—Combination of imaging systems with ancillary equipment

Definitions

  • the present invention relates to a microscope having an illumination modulus for illuminating a sample, an imaging module having a lens for imaging the sample, an excitation module exciting the sample for imaging pressure waves, and an acoustic module for dejecting the generated pressure waves.
  • the excitation module can, for example, the sample ⁇ or part of it) with a laser pulse (for example ns pulse). At least part of the introduced optical energy is absorbed by structures in the sample, which leads to a local heating and a subsequent thermoelasiic expansion and thus to a sound wave. The sound wave is detected by means of the acoustic module and can therefore serve to generate a spatially resolved image. Since typical working distances in light microscopy are usually a few mm or even sub-mm and conventional piezo transducers or hydrophones require relatively much space and at the same time an acoustic Koniakt to sample, due to these conflicting requirements, either the optical or the acoustic detection is adversely affected.
  • a laser pulse for example ns pulse
  • the object is achieved by a microscope with a lighting module for illuminating a sample, an imaging module comprising a lens for imaging the sample, an excitation module, which excites the sample for the delivery of pressure waves, and an acoustic module for detecting the generated pressure waves sample-side front lens, which defines an optically used center region, and the Akusiikmodul has an annular pressure sensor which is arranged in the region of the sample-side end of the lens and its inner diameter is selected so that, as seen in the optical axis of the objective, the optical middle range not covered.
  • the optical detection is not affected, since the optically used central area is not covered by the pressure sensor.
  • the pressure sensor can be segmented radially and / or in the circumferential direction.
  • the detection signals of the individual segments of the pressure sensor can be detected individually and then computed in a suitable manner, for example by means of a computing unit.
  • the spatial position of the virtual focus in the sample in the three spatial directions (x, y and z) can be influenced.
  • the billing can e.g. consist in first apply each individual signal with an individual time shift corresponding to a geometric delay difference and then foundedsummieren the shifted individual signals.
  • SAFT synthetic aperture focusing technique
  • this abrasion can also be done almost as fast as you like.
  • excitation and detection can be confocal with each other.
  • the pressure sensor may be attached to a damping body, which in turn is attached to the lens barrel. This can be used to reduce unwanted sound reflections on the lens barrel.
  • the pressure sensor spaced from the lens, which is understood in particular an arrangement in which there is no direct connection between the pressure sensor and lens.
  • the pressure sensor can be arranged on a cover glass or sample carrier. An arrangement on a wall in a sample chamber is also possible.
  • the lens may have a lens barrel and the pressure sensor may be attached to the lens barrel.
  • the pressure sensor may comprise a piezoceramic transducer. With such a transducer, a good sound detection is possible.
  • the pressure sensor may have an optically detectable property.
  • the objective can be designed as an immersion objective. This makes it possible that the immersion medium is also in contact with the pressure sensor, whereby a good acoustic coupling is possible.
  • the excitation module may be configured to apply electromagnetic radiation to the sample to generate pressure waves.
  • the electromagnetic radiation may, in particular, be radiation from the range of 300 nm to 3 ⁇ m, preferably 300 nm to 1300 nm, 300 nm to 1000 nm, 300 nm to 700 nm, 700 nm to 3 pm, 700 nm to 1300 nm or 700 nm to 1000 nm.
  • it is pulsed laser beams.
  • the pulse length can be in the range of ns.
  • pulses of different wavelengths can be successively used to obtain spectral information about the absorbent structures of the sample.
  • the pressure sensor may be part of the excitation module and used to generate sound waves directed at the sample.
  • the pressure sensor is used to generate pressure or sound waves and to detect the sound response coming back from the sample.
  • the microscope may include a control module that generates image data based on the data of the acoustic module.
  • the microscope according to the invention can be designed in the manner of a laser scanning-length microscope. This allows the sample to be scanned with excitation light without moving the sample.
  • the microscope according to the invention may comprise further units and modules known to the person skilled in the art for operating the microscope.
  • Fig. 1 is a schematic view of a first embodiment of the microscope according to the invention
  • Fig. 2 is an enlarged sectional view of the sample-side end of the objective 4 of Fig. 1;
  • FIG. 3 is a bottom view of the lens 4 of FIGS. 1 and 2; FIG.
  • FIG. 5 shows a view from below of the pressure sensor 9 according to a second variant
  • FIG. 6 is a bottom view of the pressure sensor 9 according to a third variant
  • Fig. 7 is a schematic sectional view of another embodiment of the microscope according to the invention.
  • Fig. 8 is a sectional view of another embodiment of the microscope according to the invention.
  • the microscope 1 according to the invention comprises a lighting module 2 for illuminating a sample 3 and an imaging module 5 having an objective 4 for imaging the sample 3.
  • the objective 4 is designed as an immersion objective. Therefore, in the schematic representation of FIG. 1 next to the sample 3, which lies between a cover glass 6 and a slide 7, an immersion medium 8 between the cover glass 6 and the cover glass 6 facing the end of the lens 4 is located.
  • the microscope 1 comprises an annular pressure sensor 9 which is arranged on the cover glass 6 or the sample 3 facing the end of the lens 4, a control module! 10 and an output unit 1.
  • the lighting module 2 can be controlled so that it pulsed electromagnetic radiation in the range of z. B. 300 nm - 3 ⁇ (hereinafter also called Anreungsstrahiung) generated via a contained in the lighting module 2 Deflection unit 12 and the lens 4 is focused in the sample 3 (for example, as a focus spot) and moved in this. Part of the energy introduced is absorbed by structures in the sample 3, which leads to a local heating and subsequent thermoelastic expansion and thus to a pressure or sound wave.
  • Anreungsstrahiung an electromagnetic radiation in the range of z. B. 300 nm - 3 ⁇
  • the sound wave is when the sample 3 z.
  • a biological sample is scattered very little in the propagation through the sample and can therefore serve to produce a spatially resolved image, with a high penetration depth in the imaging of, for example, greater than 1 mm is possible.
  • the annular pressure sensor 9 serves to detect the sound waves.
  • the objective 4 comprises a lens barrel 13 in which a plurality of lenses 14 and a sample-side front lens 15 are arranged.
  • the front lens 15 defines an optically used center region 16 which is used to apply the pulsed excitation radiation to the sample and to use it for conventional optical imaging of the sample 3 via the objective.
  • At the front end of the lens 4 of the annular pressure sensor 9 is arranged, wherein the inner diameter and the position of the pressure sensor 9 are selected so that, as seen in the direction of the optical axis 17 of the lens 4, the pressure sensor 9 does not cover the optically used central region 16.
  • the pressure sensor 9 which may also be referred to as an ultrasonic sensor, may be formed from a piezoceramic, so that a good detection of the sound waves is possible. Due to the arrangement of the pressure sensor 9 at the front end of the lens 4, the pressure sensor 9 during operation of the microscope 1 in contact with the immersion medium 8, so that a good acoustic coupling of the sample 3 to the pressure sensor 9 is present.
  • the pressure sensor 9 is connected to the control module 0, as shown schematically in FIG.
  • the control module 10 may generate image data based on the measured data of the pressure sensor 9, so that photoacoustic imaging is realized.
  • the image data can be displayed, for example, via the output unit 11.
  • the inventive arrangement of the pressure sensor 9 is no limitation of the function of the lens 4, so that a conventional light microscopy with the lens 4 is still possible. This can be used to record a preview contrast image, a fluorescence contrast image, etc.
  • the high numerical aperture of the immersion objective 4 can be used to produce a very small focus of the excitation radiation in the sample 3 for the excitation of the pressure waves. It is thus a localized excitation of the sample 3 with the pulsed one Excitation radiation (for example, laser radiation with ns pulses) possible, whereby a high spatial resolution is achieved in the photoacoustic imaging mode.
  • the excitation radiation (in particular laser radiation) generating the pressure waves can scan the sample 3 in a plane perpendicular to the optical axis 7. This can be achieved, for example, by a scanning mirror (not shown) of the deflection unit 12 arranged in the pupil of the objective 4, as is usually the case with laser scanning microscopes.
  • the pulsed excitation radiation can scan the sample 3 in the direction of the optical axis 17 by setting the focal plane of the excitation radiation accordingly. Alternatively or additionally, of course, the sample 3 can be moved accordingly.
  • the inventive arrangement of the annular pressure sensor 9 at the front end of the lens barrel 13 is a compatibility with existing microscope systems. It is only necessary to use the objective according to the invention in existing microscope systems.
  • FIG. 3 shows a view from below of the front-end of the objective 4.
  • FIG. 4 shows a variant of the pressure sensor 9 in a view (only the pressure sensor 9 being shown) from below, with the pressure sensor 9 being segmented in the axial direction in this variant.
  • the pressure sensor 9 has a plurality of concentric rings 18 (here four rings).
  • FIG. 5 shows a variant of the pressure sensor 9 in which there is a segmentation in the circumferential direction. There are several sectors 19 (here eight sectors) are provided.
  • FIG. 6 shows a variant in which the pressure sensor 9 is segmented both in the radial direction and in the circumferential direction.
  • the pressure sensor 9 may be provided with a protective cover for better cleaning or protection. This can be a plastic coating. Further, the pressure sensor 9 may not be directly attached to the lens barrel 13, but it may be arranged between the pressure sensor 9 and the lens barrel 13, a damping body (not shown). Thus, a back decoupling of the pressure sensor 9 can be realized by the lens barrel 13 to avoid sound reflections.
  • a separate connection from the control module 10 to the pressure sensor 9 is shown schematically.
  • the objective 4 can also be designed so that the electrical contacts are arranged in the objective lens.
  • the construction of the microscope according to the invention shown in FIG. 1 is only to be understood as an example. In addition to the Auflichtapt shown in Fig. 1, of course, other variants are possible. Thus, the microscope can also be designed as an inverted microscope, in which the objective 4 is arranged below the sample 3.
  • the lens 4 looks laterally into a water-filled sample chamber 20, as shown schematically in Fig. 7.
  • a seal between the lens barrel 13 and the sample chamber 20 z. B. realized by the schematically drawn annular seal 21.
  • Fig. 8 a modification of the embodiment of Fig. 7 is shown.
  • the annular pressure sensor 9 is no longer directly attached to the lens 3, but in the lens 4 facing the sample chamber wall 22.
  • the pressure sensor 9 is again arranged so that, as seen in the direction of the optical axis 17, not the optically used Center region 16 of the front lens 15 covered.
  • it is arranged in the sample chamber wall 22 so that it is in contact with the water in the sample chamber 20 or the other medium in the sample chamber 20 in order to realize the desired good Schallankoppiung.
  • the excitation of the sound waves has always been carried out optically.
  • the annular pressure sensor 9 for generating sound.
  • the control module 10 controls the pressure sensor 9 so that it sends sound waves into the sample 3 for a predetermined time and detects the sound response coming back from the sample 3.
  • the frequencies of the ultrasonic waves are, for example, 20 MHz or larger.
  • the shape of the transmitted sound waves on the relative phase delay between the individual segments of the pressure sensor 9 are changed within certain limits (eg., In the sense focusing the waves in certain depths or lateral scanning).
  • the imaging contrast is based on acoustic impendance jumps as opposed to the optical absorption contrast of the previously described photoacoustic imaging, so that these two methods provide different (possibly complementary) information about the sample 3.
  • the pressure sensor 9 can be formed from a piezoceramic. It is thus possible to use the piezoelectric effect for converting sound energy into electrical signals for pressure detection. Also, the piezoelectric effect can be used to convert electrical signals into pressure signals in the event that the pressure sensor 9 is used as a sound source.
  • any other possible type of pressure detection is possible. So z. As an annular fiber Bragg sensor or a waveguide structure for an optical detection of the ultrasonic waves are used, in this case, there is an optically detectable pressure-dependent property of the sensor, which is optically delektiert.
  • the pressure sensor can be designed as a resonant and / or broadband pressure sensor.
  • the microscope 1 according to the invention can be designed such that the excitation of the pressure waves to be detected is possible optically and / or by sound waves.

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  • Physics & Mathematics (AREA)
  • Analytical Chemistry (AREA)
  • General Physics & Mathematics (AREA)
  • Chemical & Material Sciences (AREA)
  • Optics & Photonics (AREA)
  • Biochemistry (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • General Health & Medical Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Immunology (AREA)
  • Pathology (AREA)
  • Acoustics & Sound (AREA)
  • Investigating Or Analyzing Materials By The Use Of Ultrasonic Waves (AREA)
  • Microscoopes, Condenser (AREA)

Abstract

L'invention concerne un microscope comportant un module d'éclairage (2) pour éclairer un échantillon (3), un module de reproduction (5) présentant un objectif (4) et servant à reproduire l'échantillon (3), un module d'excitation (2, 10; 9, 10) qui stimule l'échantillon (3) à produire des ondes de pression ainsi qu'un module acoustique (9, 10) pour détecter les ondes de pression produites. L'objectif (4) comprend une lentille frontale (15) située côté échantillon, qui détermine une zone médiane (16) utilisée optiquement, et le module acoustique (9, 10) présente un capteur de pression (9) annulaire qui est disposé dans la zone de l'extrémité de l'objectif (4) située côté échantillon et dont le diamètre intérieur est sélectionné de manière que, vu dans la direction de l'axe optique (17) de l'objectif (4), il ne recouvre pas la zone médiane (16) utilisée optiquement.
PCT/EP2014/053875 2013-02-28 2014-02-27 Microscope doté d'un objectif optique et acoustique combiné WO2014131853A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
DE102013203450.4A DE102013203450B4 (de) 2013-02-28 2013-02-28 Mikroskop
DE102013203450.4 2013-02-28

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WO2014131853A1 true WO2014131853A1 (fr) 2014-09-04

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WO (1) WO2014131853A1 (fr)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN110114666A (zh) * 2016-12-19 2019-08-09 赛峰集团 用于非破坏性表征材料的设备和方法

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0810450A1 (fr) * 1996-05-29 1997-12-03 Ge Yokogawa Medical Systems, Ltd. Methode et appareil d'imagerie ultrasonore
JP2006308337A (ja) * 2005-04-26 2006-11-09 Honda Electronic Co Ltd 光学顕微鏡用超音波プローブ、超音波顕微鏡システム、及び光学／超音波顕微鏡システム
WO2009055705A2 (fr) * 2007-10-25 2009-04-30 Washington University In St. Louis Microscopie photo-acoustique confocale présentant une résolution latérale optique
WO2013185784A1 (fr) * 2012-06-11 2013-12-19 Helmholtz Zentrum München Deutsches Forschungszentrum Für Gesundheit Und Umwelt (Gmbh) Système d'imagerie et procédé d'imagerie d'objet

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Publication number Priority date Publication date Assignee Title
JP2899233B2 (ja) * 1995-07-11 1999-06-02 学校法人桐蔭学園 被検体への超音波照射機能付き光学顕微鏡

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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0810450A1 (fr) * 1996-05-29 1997-12-03 Ge Yokogawa Medical Systems, Ltd. Methode et appareil d'imagerie ultrasonore
JP2006308337A (ja) * 2005-04-26 2006-11-09 Honda Electronic Co Ltd 光学顕微鏡用超音波プローブ、超音波顕微鏡システム、及び光学／超音波顕微鏡システム
WO2009055705A2 (fr) * 2007-10-25 2009-04-30 Washington University In St. Louis Microscopie photo-acoustique confocale présentant une résolution latérale optique
WO2013185784A1 (fr) * 2012-06-11 2013-12-19 Helmholtz Zentrum München Deutsches Forschungszentrum Für Gesundheit Und Umwelt (Gmbh) Système d'imagerie et procédé d'imagerie d'objet

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
ZHONGJIANG CHEN ET AL: "In vivo detection of hemoglobin oxygen saturation and carboxyhemoglobin saturation with multiwavelength photoacoustic microscopy", OPTICS LETTERS, OPTICAL SOCIETY OF AMERICA, US, vol. 37, no. 16, 15 August 2012 (2012-08-15), pages 3414 - 3416, XP001577620, ISSN: 0146-9592, [retrieved on 20120808], DOI: 10.1364/OL.37.003414 *

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN110114666A (zh) * 2016-12-19 2019-08-09 赛峰集团 用于非破坏性表征材料的设备和方法
CN110114666B (zh) * 2016-12-19 2021-12-31 赛峰集团 用于非破坏性表征材料的设备和方法

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DE102013203450B4 (de) 2017-02-09

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