EP1846739A1 - Dispositif et procede de detection d'anisotropies par microscopie optique - Google Patents

Dispositif et procede de detection d'anisotropies par microscopie optique

Info

Publication number
EP1846739A1
EP1846739A1 EP06705809A EP06705809A EP1846739A1 EP 1846739 A1 EP1846739 A1 EP 1846739A1 EP 06705809 A EP06705809 A EP 06705809A EP 06705809 A EP06705809 A EP 06705809A EP 1846739 A1 EP1846739 A1 EP 1846739A1
Authority
EP
European Patent Office
Prior art keywords
rotation
elements
influenceable
beam path
approximately
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.)
Ceased
Application number
EP06705809A
Other languages
German (de)
English (en)
Inventor
Thomas Schimming
Klaus Rink
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.)
Octax Microscience GmbH
Original Assignee
Octax Microscience 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 Octax Microscience GmbH filed Critical Octax Microscience GmbH
Publication of EP1846739A1 publication Critical patent/EP1846739A1/fr
Ceased legal-status Critical Current

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Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
    • G01N21/21—Polarisation-affecting properties
    • 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
    • G01J4/00—Measuring polarisation of light
    • G01J4/04—Polarimeters using electric detection means
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/06—Means for illuminating specimens
    • G02B21/08—Condensers
    • G02B21/14—Condensers affording illumination for phase-contrast observation

Definitions

  • the invention relates to an arrangement and a method for the microscopic optical detection of anisotropies in microscopic objects which influence the polarization properties of light.
  • the invention relates to an arrangement for the microscopy of biological cells and their anisotropies.
  • Anisotropies are usually detected by detecting changes in the properties in the optical beam path from the illumination through or through the object to be examined to an image sensor by means of elements which in turn influence the polarization properties of light and whose parameters are electrically adjustable.
  • Suitable adjustable elements are, for example, liquid crystal cells.
  • Polarization microscopy and its microbiological application have been known for a long time.
  • the change of the optical beam path by means of mechanical change is achieved by suitable elements, either manually or by motors.
  • a mechanical motion dispensing system which is preferably based on the use of voltage-variable variable optical delay liquid crystal cells, is described in US Pat. No. 5,521,705.
  • the parameter "optical delay” is usually used. This is defined as the phase shift between the x and y components of polarized light after passing through an anisotropic material, i. a material with directional refractive index.
  • the extraction of the optical delay is carried out either manually or by image processing on the basis of several microscopic images of the object to be examined in different configurations of the beam path.
  • image processing on the basis of several microscopic images of the object to be examined in different configurations of the beam path.
  • at least three images of the object in different configurations of the beam path are necessary to determine all the interesting parameters: the local light transmittance of the object and the orientation of the anisotropic property, characterized by its orientation angle in the Image plane and by their amount, usually measured in nanometers.
  • variable configuration of the beam path for isotropic objects or isotropic object areas does not have the necessary invariance. Since the isotropic regions, i. the areas of the analyzed objects which do not have anisotropies usually fill the majority of the image section, the missing invariance results in a disturbing flickering, i. the basic brightness of the microscopic image changes cyclically.
  • this considerably complicates the use of image sensors for the automatic detection and evaluation of the images of the object since the brightness fluctuations have to be subsequently corrected by computation, whereby such calculations always result in a worsened signal-to-noise ratio.
  • the use of technically conventional image sensors with adaptive brightness control is prevented, which can not follow the rapid flicker.
  • An arrangement according to the invention has a beam path which extends between a light source and an optical sensor, wherein the object is arranged in this beam path.
  • In the beam path are also an element for polarization-optical analysis and at least one influenced by a control element arranged, which causes a controllable with respect to a rotation angle rotation of the polarization properties of penetrating light.
  • a control of the arrangement has means by which at least three different angles of rotation can be predetermined and means for detecting an image of the object for each of the at least three angles of rotation by means of the optical sensor. The arrangement is such that the images of an isotropic region of the object are substantially equal in angle of rotation.
  • a light source can be provided which generates approximately monochromatic, circularly polarized light.
  • a linear polarization filter is used as the element for polarization-optical analysis.
  • an influenceable element For generating three angles of rotation, an influenceable element can be provided, in which, with appropriate control, at least three different angles of rotation can be set.
  • an element is used which allows angles of rotation of approximately 0 °, 60 ° and 120 ° or 0 °, 120 ° and 240 °.
  • two influenceable elements can also be provided, which each have two rotational angle states and are arranged so that at least three different angles of rotation can be set by controlling the two influenceable elements.
  • influenceable elements are used which have angles of rotation of 0 ° and 60 ° or 0 ° and 120 °. These elements are then arranged so that at least three different resulting rotation angles of approximately 0 °, 60 ° and 120 ° or 0 °, 120 ° and 240 ° are adjustable by influencing the two elements.
  • employable elements are used which are designed so as not to cause an optical delay or an optical delay corresponding to a multiple of the wavelength of the light generated by the light source.
  • elements are used which, in a drive-free state, cause a rotation about an angle of rotation as well as an optical delay and, in a controlled state, cause almost no rotation and almost no delay.
  • an optical compensation element for example a delay element, is arranged in the beam path whose parameters, for example delay and arrangement angle, are chosen with respect to the influenceable elements such that the images of an isotropic region of the object do not differ or only minimally differ in the different resulting rotation angles.
  • This delay element is preferably arranged in the beam path between the influenceable elements.
  • advantageously influenceable elements can be used, which are designed so that the rotation angle in the drive-free state is approximately 90 °.
  • Such elements are available inexpensively from various manufacturers and are often used as optical valves or shutters.
  • TN (Twist Nematic) liquid crystal cells are suitable as influenceable elements since these cells are also sufficiently fast for the detection with frame rates of 10-50 images per second and thus allow the detection of anisotropies of moving or moving objects.
  • the invention further relates to a method for microscopically detecting the isotropic properties of an object by means of a microscope arrangement with the following method steps: providing at least one light source; Providing at least one optical sensor;
  • An advantage of the present invention can be seen in the fact that the different (at least three) images of the object to be examined in different configurations of the beam path do not differ or only slightly differ in brightness in image regions without anisotropies, ie in isotropic image regions Image reproduction (in an eyepiece of the microscope and on the image sensor) outside the possibly existing anisotropic regions of the object to be examined is not or only minimally different from classical microscopic image reproduction.
  • Another important advantage of a system having such an invariance is the maintenance of a largely uniform temporal brightness, in particular when microscoping biological cells having large isotropic object areas.
  • This uniform basic brightness is advantageous for an automatic, time-independent adjustment of the image sensor to the illumination conditions in order to ensure optimum precision and in particular a low signal-to-noise ratio of the image pickup.
  • a uniform basic brightness is also advantageous for technically customary image sensors with adaptive brightness control, since the adaptive methods used there are designed for quasi-static, ie only slowly changing, lighting conditions.
  • the single figure shows a schematic representation of a microscope
  • a beam path 106 which extends between the light source 102 and the sensor 104.
  • An object 108 to be microscoped is usually brought into the beam path 106 by means of a specimen slide 110.
  • the microscope assembly 100 will also often include an objective 114 in practice.
  • the illumination of the object 108 to be analyzed is effected by suitably polarized light, which is immutable in time.
  • the illumination of the object 108 with circularly polarized light which is generated by inserting a circular polarizing filter 112 in the beam path between the light source 102 of the microscope and object 108.
  • a model mathematical consideration of the problem shows that (only) lighting with circular polarized light allows a complete analysis of the anisotropies independent of their orientation angle in the viewing plane.
  • An invariant arrangement for isotropic object areas which provides uniform basic brightness, is achieved by arranging in the beam path 106 between the object 108 to be analyzed and the sensor 104 one or more elements 116 which can electrically adjust the polarization properties of the light in the beam path 106 , Particularly advantageous are influenceable elements 116, which in turn thereby produce no optical delay or anisotropy. It is advantageous that those portions of the circularly polarized light from the light source 102, which were not changed by anisotropies in the object 108 in their polarization properties, which are therefore still circularly polarized, are not influenced by polarization optically measurable rotation. Therefore, a polarization-optical analysis following the beam path 106, which advantageously takes place by means of a linear polarization filter 120, will have the desired invariance properties.
  • a controller 122 shown schematically in the figure, is provided to electrically energize the element or elements 116 to adjust the optical rotation caused by element (s) 116 to a desired angle of rotation. Furthermore, it can be provided to couple the controller to the sensor 104 in order to capture an image or trigger the detection after setting a rotation angle by means of the sensor 104.
  • At least three images of the object are detected using at least three different configurations.
  • the different configurations are achieved by three different rotation angles. This may be done by one element 116A having three different rotational configurations or by two consecutive elements 116A 5 116B, each with two different rotational configurations. Of the four possible constellations resulting from two elements, three advantageous ones can be selected. If only one element 116A is provided, three different voltages corresponding to the three different angles of rotation of the element 116A are supplied by the controller 122. If, on the other hand, two elements 116A, 116B are provided, the controller supplies one of two voltage values respectively corresponding to one of the two angles of rotation to the two elements 116A, 116B for setting one of the desired three angles of rotation.
  • Suitable inexpensive elements which cause a rotation of the polarization properties of light are so-called TN (twist-nematic) liquid crystal cells whose liquid crystal in the inactive state causes a rotation of the polarization properties of light.
  • the angle of rotation can be influenced by the construction.
  • the optical delay reaches an integer multiple of the wavelength and is thus not detectable or measurable.
  • a suitable narrow wavelength band pass or interference filter can be inserted in the beam path directly after the light source 106;
  • TN liquid crystal cells By applying an electrical voltage by means of the controller 122, it is possible to convert these TN liquid crystal cells into an isotropic state in that - at least to a good approximation - neither a rotation nor an optical delay occurs. In such TN liquid cells, moreover, the transition from the inactive to the active state (and back) in a particularly short time is possible.
  • TN liquid crystal cells are much faster than the liquid crystal cells known from US Pat. No. 5,521,705 cited in the introduction, the effect of which is based on an electrical change in the optical delay and where no rotation occurs.
  • the switching times of TN liquid crystal cells allow a pseudo moving picture analysis with relevant frame rates in the order of 10-50 frames per second, which is not achievable with different liquid crystal cells due to their considerably slower electrical controllability with the necessary precision.
  • an optimal arrangement comprises two influenceable elements 116A, 116B, which can each be switched between 60 ° rotation and 0 ° rotation and are arranged in the beam path 106 one behind the other between object 108 and sensor 104 followed by an arbitrarily oriented linear polarizer 120.
  • Three configurations are used for object analysis: (a) rotation in the first element: 60 °, in the second element 60 °, (b) rotation in the first element: 60 °, in the second element 0 °, (c) rotation in the first element: 0 °, in the second element 0 °.
  • a similar effect can be achieved if in each element 116A, 116B, a rotation of 120 ° or 0 ° instead of 60 ° or 0 °.
  • Brightness information in the case of existing anisotropies is the average of the three images taken.
  • the strength of the anisotropy i. the optical local optical retardation of the object at a point in the image plane results from the difference images of (a) and (b) or (b) and (c), which are advantageously obtained by subtracting the image data in a digital image processing system (not shown) ) are obtained from the three raw images.
  • An important advantage of this exemplary arrangement is an angle-independent determination of the strength of the
  • TN liquid crystal elements with 60 ° or 120 ° rotation and vanishing optical delay in the inactive state are technically possible but not marketable and a corresponding purpose-optimized production may be too expensive.
  • TN liquid crystal elements with 90 ° rotation are customary in the market, since with such cells optical valves (so-called shutters) can be produced without problems and wavelength-independently.
  • shutters optical valves
  • even with commercially available elements with 90 ° rotation is ensured that in the electrically active state, i. after application of a sufficiently high voltage by the controller 122, both the rotation and the optical delay at least approximately disappear.
  • the optical arrangement 100 in the beam path 106 now consisting of light source 102, color filter (not shown), circular polarization filter 112th , object 108 to be analyzed, first TN liquid crystal element 116A, second TN liquid crystal element 116B, linear Polarization filter 120 and image sensor 104 are suitably compensated by an optical compensation element 118 to allow for isotropic objects or isotropic object areas three configurations with at least approximately identical image output.
  • This compensation can be achieved, for example, by inserting a suitable, preferably invariable, optical delay element 118 of suitable commercial delay into the beam path 106 between the first liquid crystal element 116A and the second liquid crystal element 116B, using as free parameters the relative orientation angles of the various ones involved in the optical analysis Components of the system 100 (first TN liquid crystal element 116A, delay element 118, second liquid crystal element 116B, linear polarization filter 120) are to be optimized.
  • compensating element 118 a compensating element or a group of elements composed of two or more elements, for example in a sandwich construction, may be provided without deviating from the present invention.
  • the arrangement of the compensation element 118 between the influenceable elements 116 represents only one of numerous possible arrangements.
  • the element may be placed elsewhere in the beam path, for example between the object and the first influenceable element 116A or between the second influenceable element 116B and the sensor 104. It is also conceivable to combine the analysis element 120 and the compensation element in one element - not shown.
  • the arrangement in accordance with the present invention may comprise further active or passive optical elements in the beam path 106, for example, the improvement of Beam path 106, the protection of the object 108 from excessive heat, magnification or image sharpness improvement can serve.
  • suitable high resolution sensors 104 are available with sufficient imaging performance, magnifying optics such as the lens 114 may be eliminated.
  • a semipermeable sensor can be provided followed by an eyepiece - not shown.
  • a prism or the like can be provided in the beam path in front of the sensor, which divides the beam path into a beam path to the sensor and a further beam path to an eyepiece - not shown.

Abstract

L'invention concerne un dispositif et un procédé de détection d'anisotropies par microscopie optique dans des objets microscopiques, influençant les caractéristiques de polarisation de la lumière. L'invention concerne notamment un dispositif présentant un couloir à rayons (106) s'étendant entre une source lumineuse (102) et un capteur optique (104), l'objet (108) étant disposé dans ce couloir à rayons (106). Ledit couloir à rayons (106) contient également un élément (120) d'analyse optique de polarisation et au moins un élément (116) pouvant être influencé par une commande (122) provoquant une rotation, commandable en ce qui concerne un angle de rotation, des caractéristiques de polarisation de la lumière traversante. La commande (122) du dispositif (100) comporte des éléments destinés à prédéterminer trois angles de rotation différents et des éléments destinés à détecter une image de l'objet (108) pour chacun des trois angles de rotation au moyen du capteur optique (104). Le dispositif (100) est conçu de telle manière que les images d'une zone isotrope de l'objet (108) sont essentiellement identiques en ce qui concerne les angles de rotation.
EP06705809A 2005-02-07 2006-01-19 Dispositif et procede de detection d'anisotropies par microscopie optique Ceased EP1846739A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
DE102005005757A DE102005005757B4 (de) 2005-02-07 2005-02-07 Anordnung zur mikroskop-optischen Erfassung von Anisotropien
PCT/DE2006/000070 WO2006081791A1 (fr) 2005-02-07 2006-01-19 Dispositif et procede de detection d'anisotropies par microscopie optique

Publications (1)

Publication Number Publication Date
EP1846739A1 true EP1846739A1 (fr) 2007-10-24

Family

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EP06705809A Ceased EP1846739A1 (fr) 2005-02-07 2006-01-19 Dispositif et procede de detection d'anisotropies par microscopie optique

Country Status (4)

Country Link
US (1) US20080170227A1 (fr)
EP (1) EP1846739A1 (fr)
DE (1) DE102005005757B4 (fr)
WO (1) WO2006081791A1 (fr)

Families Citing this family (10)

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Publication number Priority date Publication date Assignee Title
RU2419778C2 (ru) * 2005-05-25 2011-05-27 Арева Нп Способ измерения анизотропии элемента, содержащего, по меньшей мере, один делящийся материал, и соответствующая установка
DE102006051872B3 (de) * 2006-10-31 2008-07-03 Octax Microscience Gmbh Verfahren, Anordnung und Computerprogrammprodukt zur Bestimmung von Anisotropieparametern
DE102007058558A1 (de) * 2007-12-05 2009-06-25 Carl Zeiss Microimaging Gmbh Phasenkontrastmikroskop mit einer Anordnung zur Variation der Phase des Lichtes
EP2267422A1 (fr) 2009-06-26 2010-12-29 Evgeny Pavlovich Germanov Méthode et dispositif pour déterminer la malignité des matériaux biologiques
ITRM20120257A1 (it) * 2012-06-05 2013-12-06 Advanced Comp Systems A C S S P A Metodo per la quantificazione del grado di regolarità morfologica della zona pellucida in embrioni e ovociti.
DE102013106929A1 (de) * 2013-07-02 2015-01-08 JOMESA Meßsysteme GmbH Verfahren zur Analyse einer metallische und nichtmetallische Partikel enthaltenden Partikelansammlung und Vorrichtung zur Durchführung des Verfahrens
US9689793B2 (en) * 2014-02-14 2017-06-27 Kent State University System and method thereof for accurate optical detection of amphiphiles at a liquid crystal interface
KR102659810B1 (ko) * 2015-09-11 2024-04-23 삼성디스플레이 주식회사 결정화도 측정 장치 및 그 측정 방법
IT201600132813A1 (it) * 2016-12-30 2018-06-30 Istituto Naz Fisica Nucleare Metodo e apparato per rilevare particelle di dimensioni subdiffrattive
DE102021105888A1 (de) * 2021-03-11 2022-09-15 Scoobe3D Gmbh Optisches System zur Gewinnung von 3D-Rauminformationen

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US6924893B2 (en) * 2002-05-13 2005-08-02 Marine Biological Laboratory Enhancing polarized light microscopy

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Also Published As

Publication number Publication date
DE102005005757A1 (de) 2006-08-10
US20080170227A1 (en) 2008-07-17
WO2006081791A1 (fr) 2006-08-10
DE102005005757B4 (de) 2007-03-15

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