EP1203256A1 - Filtre pour microscope permettant d'ameliorer le contraste automatique - Google Patents

Filtre pour microscope permettant d'ameliorer le contraste automatique

Info

Publication number
EP1203256A1
EP1203256A1 EP00946668A EP00946668A EP1203256A1 EP 1203256 A1 EP1203256 A1 EP 1203256A1 EP 00946668 A EP00946668 A EP 00946668A EP 00946668 A EP00946668 A EP 00946668A EP 1203256 A1 EP1203256 A1 EP 1203256A1
Authority
EP
European Patent Office
Prior art keywords
image
spatial frequency
microscope
filter
value
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Withdrawn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
EP00946668A
Other languages
German (de)
English (en)
Inventor
Anders Heyden
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.)
Cellavision AB
Original Assignee
Cellavision AB
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
Priority claimed from SE9902641A external-priority patent/SE515676C2/sv
Application filed by Cellavision AB filed Critical Cellavision AB
Publication of EP1203256A1 publication Critical patent/EP1203256A1/fr
Withdrawn legal-status Critical Current

Links

Classifications

    • G—PHYSICS
    • G06—COMPUTING; CALCULATING OR COUNTING
    • G06T—IMAGE DATA PROCESSING OR GENERATION, IN GENERAL
    • G06T5/00—Image enhancement or restoration
    • G06T5/10—Image enhancement or restoration using non-spatial domain filtering
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00—Microscopes
    • G02B21/36—Microscopes arranged for photographic purposes or projection purposes or digital imaging or video purposes including associated control and data processing arrangements
    • G02B21/365—Control or image processing arrangements for digital or video microscopes
    • G—PHYSICS
    • G06—COMPUTING; CALCULATING OR COUNTING
    • G06T—IMAGE DATA PROCESSING OR GENERATION, IN GENERAL
    • G06T2207/00—Indexing scheme for image analysis or image enhancement
    • G06T2207/10—Image acquisition modality
    • G06T2207/10056—Microscopic image

Definitions

  • the present invention relates to a microscope having an image processing system and a method for such a microscope .
  • Microscopes are frequently provided with a camera for recording digital images.
  • the digital image can then be processed in a number of different ways. For example, biological preparations are studied under a microscope provided with an image processing system.
  • the image processing system can then be used to recognise a certain type of organisms or cells to be able to automatically determine the presence of the organism or cell in the specimen.
  • a standard method of improving the contrast in an image is what is referred to as the High-Boost method, in which the image is multiplied by a factor, whereupon a low-pass filtered image is subtracted from the result of the multiplication.
  • US Patent 5,696,850 discloses a method and a system using an algorithm for increasing the sharpness of reproduced images, the system having a digital camera and a reproduction device.
  • the transfer functions of the camera and the reproduction device are determined by measuring them accurately.
  • the measuring operation requires advanced and expensive apparatus and is carried out in one direction at a time since it is difficult to make a two-dimensional measurement of the transfer function.
  • the measurement of the optical transfer function must be effected each time something has been changed in the optical system.
  • the inverse Fourier transform of the two combined transfer functions is used to calculate a one-dimensional filter.
  • the filter is used to filter a recorded image by first applying the filter in one direction and subsequently in a direction at right angles to the first .
  • a further problem of the method and the device according to the US patent is that the noise at high frequencies will be amplified in an uncontrolled manner if the value of the transfer function is small for high frequencies. Moreover, a satisfactory result is not achieved with two one-dimensional filters applied successively.
  • An object of the present invention is to provide a microscope, which increases the sharpness while at the same time the noise is restricted in an image taken by means of the microscope.
  • a further object of the present invention is to provide a method for recording and processing of digital images so as to obtain sharp images .
  • a microscope with an image processing system comprises an object holder, optics which in an image plane form an image of an object placed in the image holder, and a digital image sensor, which has a number of sensor elements, for recording the image.
  • a microscope according to the invention is characterised in that the image sensor and the image plane are arranged in such manner that the spatial frequency of the sensor elements is higher than the maximum spatial frequency of the image.
  • the microscope further comprises at least one calculating means connected to the sensor, a first calculating means being adapted to provide a two-dimensional filter function, which essentially has a first value at the spatial frequency zero, a value greater than zero at a spatial frequency above the maximum spatial frequency of the image and a peak value between the spatial frequency zero and the spatial frequency of the second value, to calculate a digital filter corresponding to a two-dimensional inverse transform of the filter function, and to filter a recorded image by means of the digital filter.
  • a first calculating means being adapted to provide a two-dimensional filter function, which essentially has a first value at the spatial frequency zero, a value greater than zero at a spatial frequency above the maximum spatial frequency of the image and a peak value between the spatial frequency zero and the spatial frequency of the second value, to calculate a digital filter corresponding to a two-dimensional inverse transform of the filter function, and to filter a recorded image by means of the digital filter.
  • the filter function decreases toward zero for spatial frequencies above the spatial frequency of the peak value .
  • the first value is preferably one since no attenuation usually occurs at the spatial frequency zero. It is, however, possible to have amplification also at the spatial frequency zero in the image.
  • the spatial frequency of the sensor elements is defined as the inverse of the double distance between two adjoining sensor elements.
  • the microscope is adapted to calculate a digital filter when a user initiates such calculation.
  • the microscope is adapted to calculate a digital filter each time a new image is recorded by means of the image sensor, which, however, does not yield any particular advantages.
  • the filter function is a convolution of two one-dimensional filter functions.
  • the digital filter will then not be rotationally symmetrical .
  • the image sensor is advantageously a semiconductor sensor in the form of, for instance, a CMOS sensor or a CCD (charge coupled device) which is composed of a number of sensor elements equidistantly spaced from each other.
  • the spatial frequency of the sensor elements is the inverse of the double distance between two adjoining sensor elements .
  • the image sensor can be some other kind of image sensor such as a vidicon. What is essential for the invention is that the image sensor has a better resolution than the maximum spatial frequency of the image.
  • the sensor elements have a spatial frequency which is at least 1.5 times, advantageously at least 2 times higher, than the maximum spatial frequency of the image.
  • the microscope also comprises an input means for inputting values providing information about the filter function.
  • the microscope is adapted to make an estimate of the limit frequency of the optics by recording an image by means of the image sensor.
  • the recorded image is Fourier-transformed so that an image in the frequency plane is obtained.
  • the first calculating means calculates on the basis of the Fourier-transformed image a limit frequency.
  • the integrated signal up to the limit frequency is the major part of the total light energy of the image and advantageously at least 90%, preferably at least 95%, of the total light energy in the transformed image.
  • the limit frequency measured in the manner described is not the same as the maximum frequency which the optical system lets through, but is a usable estimate thereof .
  • the limit frequency is used to calculate the position of the peak value of the filter function. It is advantageous to let the position of the peak value depend also on the values of the appearance of the filter function, which are inputted with the aid of the input means.
  • the estimate is preferably made when a user initiates such an estimate, but alternatively the estimate is made each time a new image is recorded.
  • the filter function is advantageously strictly increasing up to the limit frequency and subsequently strictly decreasing so that the filter function up to the limit frequency conforms as far as possible with the inverse of an actual transfer function.
  • an actual transfer function is strictly decreasing up to the maximum frequency which the optical system lets through.
  • Fig. 1 is a schematic view of a microscope according to an embodiment of the present invention.
  • Fig. 2 is a block diagram of the function of a microscope according to the invention.
  • Fig. 3 illustrates the filter function as a function of the frequency in a microscope according to the present invention.
  • Fig. 4 shows a digital filter according to the present invention. Detailed Description of the Invention
  • Fig. 1 is a schematic view of a microscope according to the present invention.
  • the microscope has a source of light 1, which illuminates an object 2 placed on an object holder 3. Light from the object is collected with a microscope objective 4.
  • a digital image sensor is arranged at a distance from the microscope objective 4.
  • the digital image sensor is a CCD.
  • the CCD 5, the microscope objective 4 and the object 2 are arranged in a spaced-apart relationship so that the image plane of the microscope objective coincides with the surface of the image sensor 5.
  • the CCD is formed with a large number of sensor elements 6 which are mutually spaced apart a distance d. Each picture element corresponds to a pixel in a digital image.
  • the image sensor 5 is connected to an image processing means 7, which in turn is connected to a display 8 and an input means in the form of a keyboard 9.
  • a first calculating means 24 and a second calculating means 25 are arranged.
  • lenses 10 are arranged to transfer the image to the CCD 5.
  • the optical system consisting of the microscope objective 4 and the lenses 10 has a combined transfer function which describes how different spatial frequencies of the object 2 are transferred to the image plane. Depending on the design of the objective 4 and the lenses 10, spatial frequencies of different degrees are transferred from the object 2.
  • the maximum spatial frequency that is transferred from the object 2 to the CCD 5 is usually defined as the resolution of the optical system. Thus only structures corresponding to a certain minimum size of the object will be distinguished in the image.
  • the size of the image may vary by varying the mutual arrangement of the CCD 5, the microscope objective 4 and the object 2.
  • the spatial frequency of the sensor elements is defined as the inverse of their double mutual distance 2d.
  • the object 2, the microscope objective 4, the lenses 10 and the CCD 5 are arranged so that the spatial frequency of the sensor elements is higher than the maximum spatial frequency of the image.
  • the spatial frequency of the sensor elements is preferably 1.5 times higher than the maximum spatial frequency of the image and advantageously at least 2 times higher than the maximum spatial frequency of the image in order to obtain a reduction of noise.
  • an image will be recorded by the image sensor 5.
  • the recorded image is transferred to the image processing means 7, which processes the image before it is shown on the display 8.
  • the image processing in the image processing means 7 is affected by parameters which are inputted via the keyboard 9.
  • Fig. 2 is a block diagram of the function of the image processing means 7.
  • the recorded image from the CCD 5 is inputted at the top of the figure to block 12 and at the bottom of the figure to block 13.
  • a division into the colour components of the image is made, whereupon the divided image is transferred to block 14 for filtration by means of a digital filter.
  • a limit frequency ( ⁇ 0 ) is determined by the recorded image being transformed, whereupon co 0 is calculated as the frequency below which 95% of the integrated signal in the transformed image is present.
  • the calculation of ⁇ 0 can be carried out each time a new picture is taken or when a user of the microscope initiates a determination of ⁇ 0 .
  • a digital filter is calculated when a user initiates such a calculation via the keyboard 9.
  • Fig. 3 shows a filter function as a function of the frequency which describes an amplification as a function of the frequency.
  • parameters for the transfer function of the filter are fetched from the keyboard or from a memory 23 in the image processing means.
  • the parameters that can be fetched from the keyboard 9 in block 15 are the amplification of the filter at the limit frequency, the position of the limit frequency and the appearance of the filter function above and below the limit frequency.
  • the transfer function 17 of the filter is determined on the basis of the parameters that are fetched into block 15. The filter function 17 is determined as follows:
  • ⁇ , ⁇ and Ox are parameters which the user can modify in the input means 9.
  • the parameter ⁇ 0 can either be defined by the user or be determined automatically by the system and indicates the limit frequency.
  • the amplification at the limit frequency is determined by .
  • ⁇ and ⁇ x determine how rapidly the filter fades away at high frequencies .
  • the filter function is two-dimensional and dependent on the sum of the frequency ⁇ but independent of an angular parameter, a circular symmetrical function being obtained.
  • the filter function may have a different appearance and can, for instance, be described by one polynomial up to the limit value and another polynomial above the limit value.
  • the filter function 17 has essentially the value 1 for the frequency zero and a peak value at the limit frequency ( ⁇ 0 ) . After the limit frequency ⁇ 0 , the function decreases.
  • the filter function has, however, a value different from zero above a frequency exceeding the maximum spatial frequency of the image.
  • the limit frequency essentially conforms to the maximum spatial frequency of the image.
  • the inverse-transformed filter function is digitised and cut off to a suitable size to constitute a digital filter for the recorded image.
  • the image of which the colours have been divided is filtered two-dimensionally by means of the digital filter.
  • a voluntary grey-level transformation is made in block 19, whereupon the colours in the image are again put together in block 20.
  • the grey- level transformation is carried out if there is a risk that the grey levels have reached outside permissible values in the filtration.
  • the image is shown on the display 8 or stored in a storage means 26.
  • Fig. 4 illustrates an example of a digital filter 22 which is used in block 14. As is evident from Fig. 4, the filter is circular symmetrical around the top 21. When the image is filtered in block 14, the value in a pixel is multiplied by a value corresponding to the filter at the top 21 and added to the result of the multiplications between the adjoining pixels and a corresponding value in the filter.
  • block 12 in Fig. 2 can be omitted and the filter function be determined entirely by means of parameters that are inputted via the keyboard 9 in Fig. 1.
  • blocks 13 and 20 can be omitted if the image is not a colour image.

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  • Physics & Mathematics (AREA)
  • Engineering & Computer Science (AREA)
  • General Physics & Mathematics (AREA)
  • Multimedia (AREA)
  • Theoretical Computer Science (AREA)
  • Computer Vision & Pattern Recognition (AREA)
  • Chemical & Material Sciences (AREA)
  • Analytical Chemistry (AREA)
  • Optics & Photonics (AREA)
  • Microscoopes, Condenser (AREA)
  • Image Processing (AREA)

Abstract

Un microscope comprend un support (3) d'objet, un dispositif optique destiné, dans un plan image, à former une image d'un objet (2) placé sur le support d'objet et un détecteur (5) d'images numériques comportant plusieurs éléments (6) de détection destinés à enregistrer l'image. Le détecteur d'images et le plan image sont disposés de telle sorte que la fréquence spatiale des éléments (6) de détection soit supérieure à la fréquence spatiale maximale de l'image. Le microscope comprend en outre au moins un premier dispositif (24) de calcul connecté au détecteur (5) d'images et conçu pour fonctionner comme un filtre bidimensionnel, présentant essentiellement une valeur une à la fréquence spatiale zéro, une valeur supérieure à zéro à une fréquence spatiale supérieure à la fréquence spatiale maximale de l'image et une valeur de pointe entre ces fréquences, pour calculer un filtre numérique correspondant à une transformée de Fourier inverse bidimensionnelle de la fonction de filtre et pour filtrer une image enregistrée au moyen du filtre numérique.
EP00946668A 1999-07-09 2000-06-29 Filtre pour microscope permettant d'ameliorer le contraste automatique Withdrawn EP1203256A1 (fr)

Applications Claiming Priority (5)

Application Number Priority Date Filing Date Title
SE9902641A SE515676C2 (sv) 1999-07-09 1999-07-09 Mikroskopfilter för automatisk kontrastskärpning
SE9902641 1999-07-09
US15044099P 1999-08-24 1999-08-24
US150440P 1999-08-24
PCT/SE2000/001376 WO2001004683A1 (fr) 1999-07-09 2000-06-29 Filtre pour microscope permettant d'ameliorer le contraste automatique

Publications (1)

Publication Number Publication Date
EP1203256A1 true EP1203256A1 (fr) 2002-05-08

Family

ID=26663618

Family Applications (1)

Application Number Title Priority Date Filing Date
EP00946668A Withdrawn EP1203256A1 (fr) 1999-07-09 2000-06-29 Filtre pour microscope permettant d'ameliorer le contraste automatique

Country Status (4)

Country Link
EP (1) EP1203256A1 (fr)
JP (1) JP2003504745A (fr)
AU (1) AU6039100A (fr)
WO (1) WO2001004683A1 (fr)

Families Citing this family (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN100477734C (zh) * 2001-07-06 2009-04-08 帕朗特研究公司 成像系统以及采用倒易空间光学设计的方法
US20040184675A1 (en) * 2002-12-31 2004-09-23 Brown Carl S. Deconvolution for the reduction of blurring induced by internal reflections
US9996920B2 (en) * 2014-12-09 2018-06-12 Berkeley Lights, Inc. Automated detection and repositioning of micro-objects in microfluidic devices
CN109816609B (zh) * 2019-01-31 2021-12-21 领航基因科技（杭州）有限公司 一种基于傅里叶变换的数字pcr图像还原方法及其应用

Family Cites Families (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5561611A (en) * 1994-10-04 1996-10-01 Noran Instruments, Inc. Method and apparatus for signal restoration without knowledge of the impulse response function of the signal acquisition system
US5696850A (en) * 1995-12-21 1997-12-09 Eastman Kodak Company Automatic image sharpening in an electronic imaging system

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See references of WO0104683A1 *

Also Published As

Publication number Publication date
AU6039100A (en) 2001-01-30
WO2001004683A1 (fr) 2001-01-18
JP2003504745A (ja) 2003-02-04

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