JP2008129226A - Confocal microscope - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide a confocal microscope capable of obtaining a totally focused image whose contrast is optimumly adjusted. <P>SOLUTION: The confocal microscope 100 includes: a light condensing position varying means for varying the light condensing position of an objective lens 105 within an optional range along the optical axis direction of a converged light; a confocal image obtaining means for scanning a plane orthogonal to the optical axis direction of the converged light for every light condensing position, receiving the reflected light of the converged light from a sample 106 and obtaining the focal image; a totally focused image forming means for extracting the maximum luminous value for every pixel from the confocal image and forming the totally focused image; a height map image forming means for obtaining the height of the sample at the light condensing position based on the light condensing position where the maximum luminance value is obtained, and forming the height map image; an emphasis level control means for deciding the emphasis level of the contrast to be applied on the totally focused image in accordance with the state of the sample obtained from the height map image; and an image processing part for performing a contrast emphasis processing of the totally focused image in accordance with the decided emphasis level. <P>COPYRIGHT: (C)2008,JPO&INPIT

Description

  The present invention relates to a confocal microscope that can acquire an omnifocal image of a sample to be observed.

  The confocal microscope scans the sample surface with the light from the light source as a spot, and converts only the light reflected from the sample surface that has passed through the confocal stop into an electrical signal by the photodetector. Get 3D information.

  In a confocal microscope, the amount of light detected when focused on the optical axis is maximized and almost zero when it is out of focus, so the sample surface is moved at a predetermined pitch in the Z-axis (optical axis) direction. Then, the sample surface is two-dimensionally scanned with the spot light to obtain light quantity (maximum luminance) information that maximizes the light quantity for each pixel and position information (height information) in the Z-axis (optical axis) direction at this time. . Further, when an image is generated using only this maximum luminance information, an omnifocal image in which the entire screen is in focus is obtained.

  Therefore, in order to obtain a good omnifocal image in a confocal microscope, it is necessary to adjust the settings of the light source intensity, detector sensitivity, electrical signal offset, etc. of the device so that an optimal contrast is obtained when the image is acquired. is there.

However, in order to adjust the above settings to the optimum state while moving the sample surface in the Z-axis direction, it takes an observer's skill and effort. Also, the adjustment state is likely to vary depending on the observer.
Therefore, attempts have been made to obtain an appropriate contrast by automatically adjusting the light source intensity and the detector sensitivity from the acquired image for the image to be two-dimensionally scanned. For example, Patent Document 1 discloses a gain adjustment method that can automatically and reliably adjust the gain of a confocal microscope at a soft level in a short time.
JP 2000-321503 A

  However, in the above-described method, it is possible to optimally adjust the contrast at a specific Z position, but there is a problem that it is difficult to adjust to the optimal contrast for the omnifocal image.

  In general, methods such as histogram expansion and histogram averaging and their application methods are widely known as image processing methods for performing contrast enhancement on an acquired image.

  For example, in a confocal microscope, when materials having different reflectances exist in the field of view, they are observed as a contrast of light and shade. As described above, when a good contrast is obtained at the time of image acquisition, the enhancement effect by general contrast enhancement is processed to be small, and it is possible to adjust to an optimum contrast. Conversely, even when an image with low contrast is acquired due to insufficient light source intensity or detector sensitivity during image acquisition, the overall contrast enhancement process is performed, so the optimum contrast can be adjusted. It is.

However, when such general-purpose image processing is applied to all omnifocal images, uniform processing is performed regardless of the sample shape, and extra emphasis is applied depending on the sample.
For example, if there are minute scratches or foreign objects in a flat surface such as a mirror, contrast is necessary for that part, but it is desirable that most other surface regions can be observed as flat with low contrast. .

  However, when a general contrast enhancement process is performed, the luminance concentrated on a specific part is dispersed throughout, so that optical unevenness that was not noticeable before the process is excessively emphasized. In some cases, a good observation image could not be obtained.

  The present invention has been made in view of the above-described problems, and a problem to be solved is to provide a confocal microscope capable of acquiring an omnifocal image adjusted to an optimum contrast.

  In order to solve the above-described problems, a confocal microscope according to the present invention includes a light irradiating unit that irradiates a sample with focused light through an objective lens, and a focusing position of the objective lens with respect to the sample. Condensing position changing means for changing in an arbitrary range along the axial direction, and scanning within a predetermined plane orthogonal to the optical axis direction for each condensing position, and receiving the reflected light of the focused light from the sample A confocal image acquiring means for acquiring a confocal image, an omnifocal image generating means for extracting a maximum luminance value for each pixel from the confocal image and generating an omnifocal image, and the maximum luminance value is acquired. A height map image generating means for acquiring a height of the sample at the condensing position from the condensing position and generating a height map image, and depending on the state of the sample obtained from the height map image, Contrast applied to the focused image Comprising a enhancement level control means for determining the enhancement level, and an image processing unit for performing a contrast enhancement process on the all-focus image according enhancement level reinforcing scale level control means is determined, and.

  As described above, according to the present invention, it is possible to provide a confocal microscope that can acquire an omnifocal image adjusted to an optimum contrast.

Hereinafter, embodiments of the present invention will be described with reference to FIGS.
FIG. 1 is a diagram illustrating a configuration example of a confocal microscope 100 according to an embodiment of the present invention.
Light emitted from the light source 101 passes through the beam splitter 102 and enters the two-dimensional scanning mechanism 103. The two-dimensional scanning mechanism 103 includes a first optical scanner 103a and a second optical scanner 103b, and scans the light beam two-dimensionally and guides it to the objective lens 105.

  The light beam incident on the objective lens 105 becomes a focused light and scans the surface of the sample 106. The light reflected from the surface of the sample 106 is again introduced into the beam splitter 102 from the objective lens 105 via the two-dimensional scanning mechanism 103, then reflected by the beam splitter 102 and onto the pinhole 108 via the imaging lens 107. Condensate.

Since the pinhole 108 cuts the reflected light from other than the focal point of the sample 106, only the light passing through the pinhole 108 is detected by the photodetector 109.
The Z revolver 104 includes a plurality of objective lenses 105, and the objective lens 105 having a desired magnification can be inserted into an optical path for two-dimensional scanning. Further, it is movable in the Z-axis direction (optical axis direction), and the condensing position of the objective lens 105 and the relative position of the sample 8 (hereinafter simply referred to as “relative position”) can be changed.

The sample 106 is placed on the sample table 111 and can be moved in the XY direction (a plane direction perpendicular to the optical axis) by the stage 112.
The condensing position by the objective lens 105 is in a position conjugate with the pinhole 108. When the sample 106 is in the condensing position by the objective lens 105, the reflected light from the sample 106 is condensed on the pinhole 108. When passing through the pinhole but the sample 106 is at a position deviated from the focusing position by the objective lens 105, the reflected light from the sample 106 does not pass through the pinhole.

  Here, when the sample 106 is at the condensing position of the objective lens 105 as shown in the IZ curve showing the relationship between the relative position (Z) between the objective lens 105 and the sample 106 and the output (I) of the photodetector 109. The output of the photodetector 109 becomes maximum, and the output of the photodetector rapidly decreases as the relative position between the objective lens 105 and the sample 106 increases from this position.

  Therefore, if the two-dimensional scanning mechanism 103 is used to two-dimensionally scan the condensing point and the output of the light detector 109 is imaged in synchronization with the two-dimensional scanning mechanism 103, the sample 106 has a certain height (focus). Only the output of (position) is imaged, and an image (confocal image) obtained by optically slicing the sample 106 in a two-dimensional direction is obtained.

  Further, every time a confocal image is acquired, omnifocal image data and height map image data can be obtained by detecting the maximum luminance value and the relative position at that time for each pixel. For extracting the maximum luminance and the relative position at that time, there are a method of using the measured data as it is and a method of obtaining it by approximating the ideal curve, and either method may be used.

  Therefore, the omnifocal image data is data having at least the luminance value of each pixel of the omnifocal image as a constituent element. Further, the height map image data is data having at least a component of a focal position (hereinafter, simply referred to as “height”) at which the luminance value at each pixel of the omnifocal image is acquired.

  The omnifocal image data and height map image data obtained as described above are output to the monitor 113. The monitor 113 displays an omnifocal image and a height map image together with the operation screen.

  The computer 110 controls the two-dimensional scanning mechanism 103, the Z revolver 104, the photodetector 109, and the like to execute a program for realizing the confocal microscope 100 according to the present embodiment, and a volatile memory ( RAM), an input device (for example, a keyboard or a mouse) that is an external data input means, an output device that outputs an omnifocal image or an operation screen to the monitor 113, a program, omnifocal image data, high A storage device that stores the map image data, the level control table 110a, and the like.

  The level control table 110a that defines the contrast enhancement level according to the omnifocal image and the height map image, the processing level control means 110b that determines the contrast enhancement level based on the level control table 110a, and the omnifocal point Image processing means 110c for performing image processing such as contrast enhancement on the image.

The device user can operate each unit of the confocal microscope 100 by operating the input device according to the screen displayed on the monitor 113.
FIG. 2 is a diagram for explaining the contrast enhancement processing according to the embodiment of the present invention.

A histogram 200 shown in FIG. 2 is an example of a histogram showing the luminance distribution in the omnifocal image. The horizontal axis indicates the luminance value, and the vertical axis indicates the frequency.
The histogram 200 indicates that the image has a higher contrast as the luminance value is wider and uniformly distributed. Therefore, it is possible to adjust the contrast intensity by widening (or narrowing) the luminance distribution in the region between the lower limit level a and the upper limit level b within the dynamic range.

  In this embodiment, by changing the slope c formed by connecting the luminance value at the lower limit level a and the luminance value at the upper limit level b with a straight line, the region sandwiched between the lower limit level a and the upper limit level b is operated and contrast is adjusted. Adjust the strength of.

In addition, when changing inclination c, you may change centering on the center of a dynamic range, and you may change an average value and median value of a luminance value on an axis.
FIG. 3 shows an example of the level control table 110a according to the embodiment of the present invention.

  The level control table 301 includes a contrast “enhancement level”, a “height” of the sample 106 to be observed, and a “enhancement degree” of the contrast, and the enhancement level and the emphasis according to the height of the sample 106 to be observed. Define the emphasis level corresponding to the level.

  The value of the slope c shown in FIG. 2 is used for the enhancement degree according to the present embodiment. For example, when the enhancement degree is 1.3, the luminance distribution of the region sandwiched between the lower limit level a and the upper limit level b is expanded until the slope c is changed from 1.3 to 1.0.

Therefore, the enhancement degree 1.0 indicates that the enhancement process is not executed, and the contrast enhancement effect increases as the enhancement degree value increases.
The level control table 302 includes a contrast “enhancement level”, a surface “unevenness degree” of the sample 106 to be observed, and a contrast “enhancement degree”, depending on the surface unevenness degree of the sample 106 to be observed. Define the emphasis level and the emphasis level corresponding to the emphasis level.

  Here, FIG. 4 shows an example of an arbitrary cross section of the sample 106 and its average surface. FIG. 4 shows a cross-sectional profile d and an average surface e of the sample 106 obtained from the height map image when the horizontal axis is the pixel position of the image and the vertical axis is the height position.

  The degree of unevenness according to this example is obtained by calculating the average surface e in the cross-sectional profile d, and calculating and integrating the difference between the “height of the cross-sectional profile d” and the “height of the average surface e” for each pixel. It is obtained by dividing by the number of pixels. That is, the degree of unevenness is obtained by the following formula.

“Degree of unevenness” = Σ (“height of cross-sectional profile d” − “height of average surface e”) / “number of pixels”
... (1)
As a method for extracting the average plane, for example, a least square method is used.

  The level control table 303 uses the “deviation” used to determine the suitability of the contrast enhancement process from the “enhancement level” of contrast, the “differential value” representing the change in the cross-sectional profile d of the sample 106, and the deviation of the luminance distribution. The threshold value and the contrast “enhancement degree” are provided, and the enhancement level and the enhancement level corresponding to the enhancement level are defined according to the differential value.

The differential value is obtained by differentiating the height obtained from the height map image for each pixel. The luminance deviation is obtained from the luminance histogram of the omnifocal image.
Although the value of 1.0 or more is used for the enhancement degree described above, a value smaller than 1.0 may be used. For example, when the emphasis degree is 0.5, a process of narrowing the luminance distribution in the region sandwiched between the lower limit level a and the upper limit level b may be performed until the slope becomes 0.5 to 1.0.

Further, although the inclination c is used for the enhancement degree, the lower limit level a and the upper limit level b may be used instead of the inclination c.
Further, for the degree of unevenness, a root mean square or the like of a value obtained by calculating a difference between “actual height” and “average surface height” for each pixel may be used.

Hereinafter, a case where the level control table 301 is used will be described as a first embodiment, a case where the level control table 302 is used will be described as a second embodiment, and a case where the level control table 303 will be used will be described as a third embodiment.
(First embodiment)
FIG. 5 is a flowchart showing the process of the confocal microscope according to the first embodiment of the present invention.

When the device user performs a predetermined operation via the input device, the confocal microscope 100 starts a process of acquiring a confocal image of the sample 106.
In step S501, the confocal microscope 100 acquires the omnifocal image data and the height map image data of the sample 106 by the operation described with reference to FIG. 1, and stores them in, for example, a storage device provided in the confocal microscope 100. Then, the process proceeds to step S502.

  In step S502, the confocal microscope 100 refers to the height map image data stored in the storage device. Then, isolated point noise is removed from the height map image data. This is to prevent erroneous determination (processing) from being performed due to noise in subsequent processing.

  Here, as a method of removing isolated point noise, a method of removing noise by using a median filter is generally used, but if necessary, noise can be obtained by using a simple average filter, a threshold type average filter, or the like. May be removed.

  In step S503, the confocal microscope 100 refers to the height map image data as in step S502. Then, the maximum value and the minimum value of the height are extracted from the height map image data. Further, the difference between the maximum value and the minimum value is calculated. This difference is defined as the height of the sample 106 (hereinafter referred to as “first height”).

In step S504, the confocal microscope 100 initializes a variable n holding a level number to zero. Then, the process proceeds to step S505.
In step S505, the confocal microscope 100 refers to, for example, the level control table 301 stored at a predetermined address of the storage device, and the “height” (hereinafter, “enhancement level”) that matches the level number n = 0. , “Second height”). Then, the first height is compared with the second height.

If the second height is equal to or lower than the first height, the confocal microscope 100 proceeds to step S506.
In step S506, the confocal microscope 100 increments the level number n by 1. Then, the level number n is compared with the level number N indicating the end of the level control table 301. If the level number n is N or less, the process proceeds to step S505. If the level number n is greater than N, the process proceeds to step S509.

With the processing in steps S505 and S506, the enhancement level (level number) n corresponding to the first height (the height of the sample 106) is determined.
In step S507, the confocal microscope 100 refers to the level control table 301 stored in the storage device. Then, the “enhancement level” of the emphasis level n determined by the processes of steps S505 and S506 is acquired.

  In step S508, the confocal microscope 100 sets the enhancement degree acquired in step S507 as the inclination c. Then, as described with reference to FIG. 3, the luminance distribution of the omnifocal image data acquired in step S501 is expanded in the region between the lower limit level a and the upper limit level b so that the slope c is 1.0. Process. Thereby, contrast enhancement processing is performed.

When the above contrast enhancement processing is completed, the confocal microscope 100 proceeds to step S509.
In step S509, the confocal microscope 100 outputs the omnifocal image data to the monitor 113 to display the omnifocal image.

  As described above, the confocal microscope 100 according to the first embodiment enhances the contrast for the omnifocal image according to the height (first height) at which the omnifocal image data is acquired from the sample 106. Control the level (enhancement level) to the optimum level.

  As a result, for example, even when observing a sample with a locally concentrated luminance distribution such as a mirror surface, excessive contrast enhancement is not performed, so that a good observation image can be obtained without enhancing optical unevenness. Can be obtained.

  In addition, even when there is a height difference in the sample 106, the enhancement level (enhancement level) can be controlled to an optimum level so that sufficient contrast enhancement processing is performed on the boundary portion, so that the omnifocal image at the time of image acquisition is low. Even with contrast, it is possible to obtain an observation image on which optimum contrast enhancement has been performed.

As described above, the observer can obtain an image with optimum contrast without worrying about the state of the sample 106 being observed.
Note that the difference between the maximum value and the minimum value in the height map image data is used for the first height described above. For example, in step S501, the device user uses the omnifocal image data. When specifying the range (height range) of the sample from which the sample is acquired, the range may be used as the first height.
(Second embodiment)
FIG. 6 is a flowchart showing the process of the confocal microscope according to the second embodiment of the present invention.

When the device user performs a predetermined operation via the input device, the confocal microscope 100 starts a process of acquiring a confocal image of the sample 106.
In step S <b> 601, the confocal microscope 100 acquires the omnifocal image data and the height map image of the sample 106 by the operation described with reference to FIG. 1, and stores them in, for example, a storage device included in the confocal microscope 100. Then, the process proceeds to step S602.

  In step S602, the confocal microscope 100 refers to the height map image data stored in the storage device. Then, the isolated point noise is removed from the height map image data to prevent erroneous determination (processing) due to noise in subsequent processing. The method for removing isolated point noise is the same as in the first embodiment.

  In step S603, the confocal microscope 100 refers to the height map image data as in step S602. And the average surface e about arbitrary cross-sectional profiles d in the sample 106 is extracted from the height map image data. In the present embodiment, as shown in FIG. 4, the average surface e is extracted using the least square method, but the average surface e may be obtained using an nth-order spline interpolation function or the like.

  In step S604, the confocal microscope 100, as described with reference to FIG. 4, substitutes the height of the cross-sectional profile d and the height of the average surface e obtained in step S603 into the equation (1) to obtain the degree of unevenness (hereinafter, (Referred to as “first unevenness degree”).

In step S605, the confocal microscope 100 initializes a variable n holding a level number to 0. Then, the process proceeds to step S606.
In step S606, the confocal microscope 100 refers to, for example, the level control table 302 stored at a predetermined address of the storage device, and the “concave / convex degree” of the “enhancement level” that matches the level number n = 0 (hereinafter referred to as “unevenness degree”). , “Second unevenness degree”). Then, the first unevenness degree and the second unevenness degree are compared.

If the second unevenness degree is equal to or less than the first unevenness degree, the confocal microscope 100 moves the process to step S607.
In step S607, the confocal microscope 100 increments the level number n by 1. Then, the level number n is compared with the level number N indicating the end of the level control table 302. If the level number n is N or less, the process proceeds to step S606. If the level number n is greater than N, the process proceeds to step S610.

By the processing in steps S606 and S607, the enhancement level (level number) n corresponding to the first degree of unevenness is determined.
In step S608, the confocal microscope 100 refers to the level control table 302 stored in the storage device. Then, the “enhancement level” of the emphasis level n determined by the processes of steps S606 and S607 is acquired.

  In step S609, the confocal microscope 100 sets the enhancement degree acquired in step S608 as the inclination c. Then, as described in FIG. 3, the luminance distribution of the omnifocal image data acquired in step S601 is widened so that the slope c is 1.0, and the luminance distribution in the region between the lower limit level a and the upper limit level b is widened. The contrast enhancement process is performed.

  When the above contrast enhancement processing is completed, the confocal microscope 100 proceeds to step S610. Then, the confocal microscope 100 outputs the omnifocal image data to the monitor 113 and displays the omnifocal image.

  As described above, the confocal microscope 100 according to the second embodiment optimizes the contrast enhancement level (enhancement level) for the omnifocal image according to the degree of unevenness in an arbitrary cross section (surface) of the sample 106. Control to the right level.

  As a result, as in the first embodiment, even when observing a sample with a locally concentrated luminance distribution such as a mirror surface, excessive contrast enhancement is suppressed and optical unevenness is not enhanced. An observation image can be obtained.

  Even when there is a height difference in the sample 106, the enhancement level (enhancement level) can be controlled to an optimum level so that sufficient contrast enhancement processing is performed on the boundary portion, so that the omnifocal image at the time of image acquisition is low. Even with contrast, it is possible to obtain an observation image on which optimum contrast enhancement has been performed.

  Further, the confocal microscope 100 according to the second embodiment controls the contrast enhancement level (enhancement degree) for the all-focus image to an optimum level according to the unevenness degree in an arbitrary cross section (surface) of the sample 106. Therefore, even when omnifocal image data is acquired for the tilted sample 106 as shown in FIG. 4, it is possible to obtain an omnifocal image with optimum contrast enhancement.

As described above, the observer can obtain an image with optimum contrast without worrying about the state of the sample 106 being observed.
(Third embodiment)
FIG. 7 is a flowchart showing the process of the confocal microscope according to the third embodiment of the present invention.

When the device user performs a predetermined operation via the input device, the confocal microscope 100 starts a process of acquiring a confocal image of the sample 106.
In step S <b> 701, the confocal microscope 100 acquires the omnifocal image data and the height map image of the sample 106 by the operation described with reference to FIG. 1, and stores them in, for example, a storage device included in the confocal microscope 100. Then, the process proceeds to step S702.

  In step S702, the confocal microscope 100 refers to the height map image data stored in the storage device. Then, the isolated point noise is removed from the height map image data to prevent erroneous determination (processing) due to noise in subsequent processing. The method for removing isolated point noise is the same as in the first embodiment.

  In step S703, the confocal microscope 100 refers to the height map image data as in step S702. Then, height data for an arbitrary cross-sectional profile d in the sample 106 is acquired from the height map image data. And the differential value of the height data in each pixel is calculated.

  In step S704, the confocal microscope 100 compares the differential value calculated in step S703 with a predetermined threshold M. Then, the sum of the squares (hereinafter referred to as “first differential value”) is obtained for the differential value equal to or greater than the threshold value M.

  In addition, although the process of step S703 and S704 may be performed about the whole surface of the sample 106, in a present Example, in order to reduce a calculation amount, it may be performed about the partial area | region (arbitrary cross-sectional profile d) of the sample 106. Yes. In step S704, the sum of values obtained by dividing the differential value in each pixel by the luminance value of the pixel may be used instead of the sum of the squares.

In step S705, the confocal microscope 100 initializes a variable n holding a level number to 0. Then, the process proceeds to step S706.
In step S706, the confocal microscope 100 refers to, for example, the level control table 303 stored at a predetermined address of the storage device, and the “differential value” (hereinafter, “enhancement level”) that matches the level number n = 0. , "Second differential value"). Then, the first differential value is compared with the second differential value.

If the second differential value is less than or equal to the first differential value, the confocal microscope 100 moves the process to step S707.
In step S707, the confocal microscope 100 increments the level number n by 1. Then, the level number n is compared with the level number N indicating the end of the level control table 303. If the level number n is N or less, the process proceeds to step S706. If the level number n is greater than N, the process proceeds to step S712.

The enhancement level (level number) n corresponding to the first differential value is determined by the processing in steps S706 and S707.
In step S708, the confocal microscope 100 obtains the luminance deviation σ from the luminance histogram of the omnifocal image data acquired in step S701. Then, the process proceeds to step S709.

  In step S709, the confocal microscope 100 refers to the level control table 303 as in step S706, and acquires a deviation threshold value that matches the level number n. Then, the brightness deviation σ obtained in step S708 is compared with the deviation threshold.

If the luminance deviation σ is equal to or smaller than the deviation threshold, the process proceeds to step S712. If the luminance deviation σ is larger than the deviation threshold, the process proceeds to step S710.
In step S710, the confocal microscope 100 refers to the level control table 303 stored in the storage device. Then, the “enhancement level” of the emphasis level n determined by the processes of steps S706 and S707 is acquired.

  In step S711, the confocal microscope 100 sets the enhancement degree acquired in step S10 as the inclination c. Then, as described with reference to FIG. 3, the luminance distribution of the omnifocal image data acquired in step S701 is expanded in the region between the lower limit level a and the upper limit level b so that the slope c is 1.0. The contrast enhancement process is performed.

  When the above contrast enhancement processing is completed, the confocal microscope 100 proceeds to step S712. Then, the confocal microscope 100 outputs the omnifocal image data to the monitor 113 and displays the omnifocal image.

  As described above, the confocal microscope 100 according to the third embodiment optimizes the contrast enhancement level (enhancement degree) for the omnifocal image in accordance with the differential value in an arbitrary cross section (surface) of the sample 106. Control to the right level.

  As a result, as in the first embodiment, even when observing a sample with a locally concentrated luminance distribution, such as a mirror surface, it is possible to suppress excessive contrast enhancement and provide excellent observation without optical unevenness such as shading. An image can be obtained.

  Even when there is a height difference in the sample 106, the enhancement level (enhancement level) can be controlled to an optimum level so that sufficient contrast enhancement processing is performed on the boundary portion, so that the omnifocal image at the time of image acquisition is low. Even with contrast, it is possible to obtain an observation image on which optimum contrast enhancement has been performed.

  Further, when the luminance deviation σ is equal to or less than the deviation threshold (when it is determined that the surface is a mirror or the like), the contrast enhancement process is not performed, and thus an optical such as shading caused by excessive contrast enhancement on the surface of the mirror or the like. Unevenness can be reliably prevented.

As described above, the observer can obtain an image with optimum contrast without worrying about the state of the sample 106 being observed.
In the first and second embodiments, an example is shown in which the contrast enhancement level (enhancement level) for the omnifocal image is controlled to an optimum level in accordance with “height” and “degree of unevenness”, respectively. However, as in the third embodiment, a “deviation threshold” may be used in combination.

  For example, by using a level control table of a combination of “height” and “deviation threshold” in the first embodiment and “roughness degree” and “deviation threshold” in the second embodiment, the contrast of the omnifocal image The enhancement level (enhancement level) may be controlled to an optimum level.

In this case, the processes of steps S708 and S709 may be added to the flowcharts shown in FIGS.
In the first to third embodiments, a case is described in which contrast enhancement processing is directly performed on the omnifocal image data acquired in step S501 (S601, S701) (in this case, the omnifocal image data itself is changed). However, display data in which contrast enhancement is applied to the omnifocal image data may be newly generated and output to the monitor 113 without changing the omnifocal image data itself.

  In this case, since the omnifocal image data maintains the raw data at the time of acquisition, the observer can observe the image with good contrast and perform image processing, image measurement, and analysis based on the raw data. Can be applied.

It is a figure which shows the structural example of the confocal microscope which concerns on the Example of this invention. It is a figure explaining the contrast emphasis processing concerning the example of the present invention. The example of the level control table which concerns on the Example of this invention is shown. It is a figure which shows the example of the height map image about the arbitrary cross sections in a sample, and its average surface. It is a flowchart which shows the process of the confocal microscope which concerns on 1st Example of this invention. It is a flowchart which shows the process of the confocal microscope which concerns on the 2nd Example of this invention. It is a flowchart which shows the process of the confocal microscope which concerns on the 3rd Example of this invention.

Explanation of symbols

DESCRIPTION OF SYMBOLS 100 ... Confocal microscope 101 ... Light source 102 ... Beam splitter 103 ... Two-dimensional scanning mechanism 104 ... Z revolver 105 ... Objective lens 106 ... Sample 107 ... Imaging lens 108 ... Pinhole 109 ... Photodetector 110 ... Computer 111 ... Sample stage 112 ... Stage 113 ... Monitor 200 ... Histogram 301-303 showing luminance distribution ... Level Control table

Claims (6)

A light irradiation means for irradiating the sample with focused light through an objective lens;
A condensing position changing means for changing the condensing position of the objective lens with respect to the sample in an arbitrary range along the optical axis direction of the focused light;
A confocal image acquisition unit that scans a predetermined plane orthogonal to the optical axis direction for each condensing position, receives reflected light from the sample of the focused light, and acquires a confocal image;
Omnifocal image generation means for extracting a maximum luminance value for each pixel from the confocal image and generating an omnifocal image;
A height map image generating means for acquiring a height of the sample at the condensing position from the condensing position where the maximum luminance value is acquired, and generating a height map image;
An emphasis level control means for determining an emphasis level of contrast applied to the omnifocal image according to the state of the sample obtained from the height map image;
An image processing unit that performs contrast enhancement processing on the omnifocal image according to the enhancement level determined by the enhancement level control means;
A confocal microscope characterized by comprising: Level control information storage means for storing level control information that defines the enhancement level according to the state of the sample;
The enhancement level control means reads and references the level control information from the level control information storage means, and obtains an enhancement level corresponding to the state of the sample obtained from the height map image.
The confocal microscope according to claim 1.   The confocal microscope according to claim 2, wherein a difference between a maximum value and a minimum value of the sample height obtained from the height map image is used as the sample state.   The confocal microscope according to claim 2, wherein the state of the sample uses a degree of unevenness of an arbitrary surface of the sample obtained from the height map image.   The confocal microscope according to claim 2, wherein a change rate of an arbitrary surface of the sample obtained from the height map image is used for the state of the sample.   The enhancement level control means calculates a luminance deviation from the omnifocal image, compares the luminance deviation with a preset deviation threshold value, and emphasizes from the level control information only when the luminance deviation is larger than the deviation threshold value. The confocal microscope according to claim 2, wherein a level is acquired and the image processing unit is caused to execute contrast enhancement processing.