DE4419940A1 - Three=dimensional image recording for confocal light microscopy - Google Patents

Three=dimensional image recording for confocal light microscopy

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Publication number
DE4419940A1
DE4419940A1 DE19944419940 DE4419940A DE4419940A1 DE 4419940 A1 DE4419940 A1 DE 4419940A1 DE 19944419940 DE19944419940 DE 19944419940 DE 4419940 A DE4419940 A DE 4419940A DE 4419940 A1 DE4419940 A1 DE 4419940A1
Authority
DE
Germany
Prior art keywords
light
light microscopy
light source
pinhole
confocal
Prior art date
Legal status (The legal status 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 status listed.)
Withdrawn
Application number
DE19944419940
Other languages
German (de)
Inventor
Eberhard Dipl Phys Tuengler
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Individual
Original Assignee
Individual
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 Individual filed Critical Individual
Priority to DE19944419940 priority Critical patent/DE4419940A1/en
Publication of DE4419940A1 publication Critical patent/DE4419940A1/en
Withdrawn legal-status Critical Current

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Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01B—MEASURING LENGTH, THICKNESS OR SIMILAR LINEAR DIMENSIONS; MEASURING ANGLES; MEASURING AREAS; MEASURING IRREGULARITIES OF SURFACES OR CONTOURS
    • G01B11/00—Measuring arrangements characterised by the use of optical techniques
    • G01B11/24—Measuring arrangements characterised by the use of optical techniques for measuring contours or curvatures
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01J—MEASUREMENT OF INTENSITY, VELOCITY, SPECTRAL CONTENT, POLARISATION, PHASE OR PULSE CHARACTERISTICS OF INFRARED, VISIBLE OR ULTRAVIOLET LIGHT; COLORIMETRY; RADIATION PYROMETRY
    • G01J3/00—Spectrometry; Spectrophotometry; Monochromators; Measuring colours
    • G01J3/28—Investigating the spectrum
    • G01J3/2823—Imaging spectrometer
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/0004—Microscopes specially adapted for specific applications
    • G02B21/002—Scanning microscopes
    • G02B21/0024—Confocal scanning microscopes (CSOMs) or confocal "macroscopes"; Accessories which are not restricted to use with CSOMs, e.g. sample holders
    • G02B21/0052—Optical details of the image generation
    • G02B21/0064—Optical details of the image generation multi-spectral or wavelength-selective arrangements, e.g. wavelength fan-out, chromatic profiling
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/18—Arrangements with more than one light path, e.g. for comparing two specimens
    • G02B21/20—Binocular arrangements
    • G02B21/22—Stereoscopic arrangements
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01B—MEASURING LENGTH, THICKNESS OR SIMILAR LINEAR DIMENSIONS; MEASURING ANGLES; MEASURING AREAS; MEASURING IRREGULARITIES OF SURFACES OR CONTOURS
    • G01B2210/00—Aspects not specifically covered by any group under G01B, e.g. of wheel alignment, caliper-like sensors
    • G01B2210/50—Using chromatic effects to achieve wavelength-dependent depth resolution

Abstract

The light passing through the apertured partition (8), is detected by a detector system (9,10,11). An objective lens (4) with strong chromatic aberration is used, and the light coming through the apertured partition (8) is spectrally split up by a prism (10). Several detectors (11) are provided, with which the intensity is established for each wavelength range. A light source (1) with several wavelengths or a light source with a partially continuous spectrum is used to illuminate an object (5), where e.g. two points (6,7) represent different reflected wavelengths.

Description

Die Erfindung bezieht sich auf ein 3D-Bilderkennungsverfahren mit konfokaler Licht­ mikroskopie nach dem Oberbegriff des Patentanspruchs 1.The invention relates to a 3D image recognition method with confocal light Microscopy according to the preamble of claim 1.

Das heute verwendete Verfahren zur konfokalen Lichtmikroskopie tastet jeden Objektpunkt (5) mit Hilfe eines Scanners (3) einzeln nacheinander ab und mißt die von jedem Objektpunkt (5) rückgestreute Intensität.The method used today for confocal light microscopy scans each object point ( 5 ) individually with the aid of a scanner ( 3 ) and measures the intensity backscattered from each object point ( 5 ).

Aufgabe der Erfindung ist es, ein 3D-Bilderkennungsverfahren mit konfokaler Lichtmikroskopie anzugeben, das in der Lage ist, das Bild dreidimensional zu erfas­ sen.The object of the invention is a 3D image recognition method with confocal Specify light microscopy that is able to capture the image three-dimensionally sen.

Die Erfindung wird nun anhand eines Ausführungsbeispiels in Verbindung mit der Zeichnung näher erläutert. Die Zeichnung enthält eine schematische Anordnung der Einzelelemente.The invention will now be described using an exemplary embodiment in conjunction with the drawing explained in more detail. The drawing contains a schematic arrangement of the individual elements.

Die Anordnung besteht im Wesentlichen aus einer Lichtquelle (1), wobei erfin­ dungsgemäß eine Lichtquelle mit mehreren Wellenlängen oder teilweise kontinuier­ lichem Spektrum verwendet wird.The arrangement essentially consists of a light source ( 1 ), a light source having a plurality of wavelengths or a partially continuous spectrum being used in accordance with the invention.

Nun wird, wie üblich, das Licht durch einen Strahlteiler (2) und einen Scan­ ner (3) geführt und gelangt dann durch ein schematisch angedeutetes Linsensystem (4) zum Objekt (5). Da erfindungsgemäß eine Objektivlinse (4) mit chromatischer Aberration verwendet wird, bedeutet dies, daß die Brennpunkte für verschiedenen Wellenlängen in verschiedenen Tiefen liegen. In der Zeichnung sind zwei Brenn­ punkte (6) und (7) für zwei verschiedene Wellenlängen angedeutet.Now, as usual, the light is passed through a beam splitter ( 2 ) and a scan ner ( 3 ) and then reaches the object ( 5 ) through a schematically indicated lens system ( 4 ). Since an objective lens ( 4 ) with chromatic aberration is used according to the invention, this means that the focal points for different wavelengths are at different depths. In the drawing, two focal points ( 6 ) and ( 7 ) are indicated for two different wavelengths.

Das vom Objekt (5) an den verschiedenen Brennpunkten (6) (7) rückgestreute Licht gelangt dann durch den Strahlteiler (2) auf die Lochblende (8), die die in der konfokalen Lichtmikroskopie übliche Funktion der Ausblendung des von anderen Objektpunkten rückgestreuten Lichtes besitzt.The light backscattered by the object ( 5 ) at the various focal points ( 6 ) ( 7 ) then passes through the beam splitter ( 2 ) to the pinhole ( 8 ), which has the function of blanking out the backscattered light from other object points, as is customary in confocal light microscopy .

Erfindungsgemäß wird nun das von der Lochblende kommende Licht gesammelt (9) und spektral mit (beispielsweise) einem Prisma (10) zerlegt.According to the invention, the light coming from the pinhole is now collected ( 9 ) and spectrally broken down with (for example) a prism ( 10 ).

Mit dem Detektorsystem wird die Intensität der einzelnen Wellenlängenbereiche gemessen. Jeder Wellenlänge entspricht aber ein verschieden tief liegender Brenn­ punkt. Damit kann also durch eine nachgeschaltete Verarbeitungsstufe das Bild drei dimensional erfaßt werden.With the detector system, the intensity of the individual wavelength ranges measured. However, each wavelength corresponds to a differently located burner Point. This means that the image can be processed through a subsequent processing stage three dimensionally.

Falls die Intensität des eingestrahlten Lichtes stark genug gewählt werden kann oder die Empfindlichkeit des Detektorsystems hoch genug ist, besteht ein Vorteil des Verfahrens in der im Vergleich zum zweidimensionalen Aufbau gleich großen Erfassungsgeschwindigkeit der Meßdaten.If the intensity of the incident light can be chosen strong enough or the sensitivity of the detector system is high enough, there is an advantage of the process in the same size compared to the two-dimensional structure Acquisition speed of the measurement data.

Bemerkung: Mit Ausnahme der Objektivlinse haben alle anderen Linsensysteme eine sehr kleine chromatische Aberration. Note: With the exception of the objective lens, all other lens systems have a very small chromatic aberration.  

Im Rahmen der Erfindung sind folgende Abwandlungen möglich:
Der verwendete Wellenlängenbereich kann so gewählt werden, daß die Wellenlängen­ abhängigkeit der Reflexion in guter Näherung konstant ist. Wird ein sehr schmaler Wellenlängenbereich gewählt, muß die chromatische Aberration der Objektivlinse stark sein und die spektrale Auflösung des Detektorsystems (9) (10) (11) hoch. Alter­ nativ kann ein breiter Wellenlängenbereich, eine schwache chromatische Aberration der Objektivlinse (4) und ein kleineres Auflösungsvermögen des Detektorsystems (11) gewählt werden, falls die verwendeten Objekte nur eine schwache Wellenlängen­ abhängigkeit der Reflexion aufweisen.The following modifications are possible within the scope of the invention:
The wavelength range used can be chosen so that the wavelength dependence of the reflection is constant to a good approximation. If a very narrow wavelength range is selected, the chromatic aberration of the objective lens must be strong and the spectral resolution of the detector system ( 9 ) ( 10 ) ( 11 ) high. Alternatively, a broad wavelength range, a weak chromatic aberration of the objective lens ( 4 ) and a smaller resolution of the detector system ( 11 ) can be selected if the objects used have only a weak wavelength dependence on the reflection.

Statt des angedeuteten Prismas können hier verschiedene andere zur spektralen Zerlegung geeignetere Anordnungen (beispielsweise Strichgitter, Beugungsgitter etc.) verwendet werden, die ein genügend gutes Auflösungsvermögen besitzen.Instead of the indicated prism, various others can be used for the spectral one Disassembly of more suitable arrangements (e.g. grating, diffraction grating, etc.) can be used, which have a sufficiently good resolution.

Claims (2)

1. 3D-Bilderkennungsverfahren mit konfokaler Lichtmikroskopie, bestehend aus ei­ nem in der konfokalen Lichtmikroskopie üblichen Mikroskop, bei welchem das vom Objektpunkt (5) rückgestreute Licht auf eine Lochblende (8) abgebildet wird und das durch die Lochblende (8) tretende Licht voll einem Detektorsystem (9) (10) (11) detektiert wird, dadurch gekennzeichnet, daß eine Objektivlinse (4) mit starker chromatischer Aberration verwendet wird, und das vom Objekt (5) reflektierte und durch die Lochblende (4) gekommene Licht spektral zerlegt wird, und dann mit mehreren Detektoren (11) für jeden Wellenlängenbereich die Intensität festgestellt wird.1. 3D image recognition method using confocal light microscopy, consisting of a microscope customary in confocal light microscopy, in which the light scattered back from the object point ( 5 ) is imaged on a pinhole ( 8 ) and the light passing through the pinhole ( 8 ) is fully one Detector system ( 9 ) ( 10 ) ( 11 ) is detected, characterized in that an objective lens ( 4 ) with strong chromatic aberration is used and the light reflected from the object ( 5 ) and coming through the pinhole ( 4 ) is spectrally broken down, and then using several detectors ( 11 ) to determine the intensity for each wavelength range. 2. Verfahren nach Anspruch 1, dadurch gekennzeichnet, daß eine Lichtquelle (1) mit mehreren Wellenlängen oder eine Lichtquelle mit teilweise kontinuierlichem Spektrum verwendet wird.2. The method according to claim 1, characterized in that a light source ( 1 ) with several wavelengths or a light source with a partially continuous spectrum is used.
DE19944419940 1994-06-08 1994-06-08 Three=dimensional image recording for confocal light microscopy Withdrawn DE4419940A1 (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
DE19944419940 DE4419940A1 (en) 1994-06-08 1994-06-08 Three=dimensional image recording for confocal light microscopy

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
DE19944419940 DE4419940A1 (en) 1994-06-08 1994-06-08 Three=dimensional image recording for confocal light microscopy

Publications (1)

Publication Number Publication Date
DE4419940A1 true DE4419940A1 (en) 1995-12-14

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Family Applications (1)

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DE19944419940 Withdrawn DE4419940A1 (en) 1994-06-08 1994-06-08 Three=dimensional image recording for confocal light microscopy

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DE (1) DE4419940A1 (en)

Cited By (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
DE19612846A1 (en) * 1996-03-30 1997-10-02 Zeiss Carl Jena Gmbh Arrangement for producing defined coloured longitudinal defect in confocal microscopic beam path
WO1998044375A2 (en) * 1997-03-29 1998-10-08 Carl Zeiss Jena Gmbh Confocal microscopic device
WO1999039231A1 (en) * 1998-01-28 1999-08-05 Leica Microsystems Heidelberg Gmbh Device for simultaneously detecting several spectral ranges of a light beam
GB2344014A (en) * 1998-11-19 2000-05-24 Medical Res Council Scanning confocal optical microscope system
WO2007124858A1 (en) 2006-04-26 2007-11-08 Carl Zeiss Microimaging Gmbh Microscope and microscope microexamination procedure method for the measurement of the surface profile of an object
EP2305095A1 (en) * 2009-09-30 2011-04-06 Fujifilm Corporation Imaging apparatus
CN102188290A (en) * 2010-03-19 2011-09-21 卡尔斯特里姆保健公司 3-D imaging using telecentric defocus
WO2011131311A1 (en) * 2010-04-19 2011-10-27 Witec Wissenschaftliche Instrumente Und Technologie Gmbh Apparatus for imaging a sample surface
WO2022099929A1 (en) * 2020-11-16 2022-05-19 海伯森技术（深圳）有限公司 Line spectral confocal sensor

Cited By (16)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
DE19612846C2 (en) * 1996-03-30 2000-04-20 Zeiss Carl Jena Gmbh Arrangement for generating a defined longitudinal color error in a confocal microscopic beam path
DE19612846A1 (en) * 1996-03-30 1997-10-02 Zeiss Carl Jena Gmbh Arrangement for producing defined coloured longitudinal defect in confocal microscopic beam path
US6674572B1 (en) 1997-03-29 2004-01-06 Carl Zeiss Jena Gmbh Confocal microscopic device
WO1998044375A3 (en) * 1997-03-29 1999-03-04 Zeiss Carl Jena Gmbh Confocal microscopic device
WO1998044375A2 (en) * 1997-03-29 1998-10-08 Carl Zeiss Jena Gmbh Confocal microscopic device
WO1999039231A1 (en) * 1998-01-28 1999-08-05 Leica Microsystems Heidelberg Gmbh Device for simultaneously detecting several spectral ranges of a light beam
US6614526B1 (en) 1998-01-28 2003-09-02 Leica Microsystems Heidelberg Gmbh Device for simultaneously detecting several spectral ranges of a light beam
GB2344014A (en) * 1998-11-19 2000-05-24 Medical Res Council Scanning confocal optical microscope system
GB2344014B (en) * 1998-11-19 2001-02-07 Medical Res Council Scanning confocal optical microscope system
WO2007124858A1 (en) 2006-04-26 2007-11-08 Carl Zeiss Microimaging Gmbh Microscope and microscope microexamination procedure method for the measurement of the surface profile of an object
US7715021B2 (en) 2006-04-26 2010-05-11 Carl Zeiss Microimaging Gmbh Microscope and microscope microexamination procedure method for the measurement of the surface profile of an object
EP2305095A1 (en) * 2009-09-30 2011-04-06 Fujifilm Corporation Imaging apparatus
CN102188290A (en) * 2010-03-19 2011-09-21 卡尔斯特里姆保健公司 3-D imaging using telecentric defocus
WO2011131311A1 (en) * 2010-04-19 2011-10-27 Witec Wissenschaftliche Instrumente Und Technologie Gmbh Apparatus for imaging a sample surface
US9891418B2 (en) 2010-04-19 2018-02-13 Witec Wissenschaftliche Instrumente Und Technologie Gmbh Apparatus for imaging a sample surface
WO2022099929A1 (en) * 2020-11-16 2022-05-19 海伯森技术（深圳）有限公司 Line spectral confocal sensor

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