JP2000321503A - Automatic adjusting method for confocal microscope and confocal microscope - Google Patents

Abstract

PROBLEM TO BE SOLVED: To eliminate the need for a mass storage memory and to prevent the lowering of working efficiency by surely automatically adjusting a gain only at a soft level in a short time by automatically adjusting the gain of a photodetector from the information of one image plane by a retrieval technique. SOLUTION: The gain of the photodetector 19b is automatically adjusted from the information of one image plane by the retrieval technique. Namely, a CPU 81 arithmetically calculates the optimum gain of a variable gain amplifier circuit 46 by the retrieval technique of the optimum gain in order to make an output value stored in a frame memory 84 or a value obtained from that value coincide with a set target value stored in a target value memory 88. Then, the circuit 46 is set through a variable gain D/A by the gain obtained by arithmetic calculation, and the light receiving signal of the photodetector 19b is amplified by the gain, and the output value A/D converted by an A/C conversion circuit 41 is compared with the set target value. When they do not coincide with each other, the output value is made close to the set target value by repeating the arithmetic calculation and the comparison by the CPU 81.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a microscope using the confocal principle, and more particularly to a method for adjusting a gain of a light receiving signal of a light receiving element which receives a light amount returned from a work.

[0002]

2. Description of the Related Art A microscope using the confocal principle is provided with an objective lens and a pinhole. When a sample is located at a focal position of the objective lens, a laser beam passing through the pinhole is used as a first light receiving element. , The image of the portion at the height desired to be observed, that is, only the confocal image is clearly displayed, so that the resolution is increased. As described above, the confocal optical system has a feature that the luminance is maximized at the focused position. Therefore, the sample is moved in the Z-axis direction, and the maximum luminance signal is stored in each pixel of the image. Eventually, an in-focus image is obtained.

However, since such a confocal image is a black-and-white (achromatic) image, the present applicant obtains a color image by using color information from the second light receiving element for the black-and-white image. We have invented and filed a compact and low-cost color (chromatic) confocal microscope that can be made (Japanese Patent Application No. Hei 9
-180647). According to this, by adopting a confocal optical system using a He-Ne laser as a light source,
High-resolution images beyond the limits of ordinary optical microscopes
The image can be obtained from the light receiving element, and a color image can be obtained because the color information from the second light receiving element is used. As a result, it is possible to observe the scratches and attached matter in detail.

Hereinafter, this color confocal microscope which is the basis of the present invention will be described with reference to FIGS.
In FIG. 5, the microscope is a laser optical system (first optical system).
1 and a white light optical system (second optical system) 2.
The laser optical system 1 is a confocal optical system that can detect information on the depth of the sample w, and uses, for example, a He-Ne laser 10 that emits red laser light L1 as a light source. On the optical axis of the laser 10, a first collimating lens 11, a polarizing beam splitter 12, a quarter-wave plate 13, a two-dimensional scanning device 14, a first relay lens 15, a second relay lens 1
6 and an objective lens 17 are arranged in this order. A sample stage 30 is provided near the focal position of the objective lens 17, and the objective lens 17 transmits the laser light L1 to the sample w.
Focus on the surface of. The two-dimensional scanning device 14 includes, for example, an acousto-optic deflecting element in the horizontal direction and a galvanomirror in the vertical direction, and deflects the laser beam L1 to obtain the sample w.
The light condensing position on the surface is set two-dimensionally (X, Y) along the surface of the sample w.
Direction) in real time. Sample stage 30
Is driven and controlled in the Z (vertical) direction by a stage control circuit 40, and can be moved by a manual handle in the X and Y directions.

[0005] The laser light (response light) L reflected by the sample w
1 passes through the objective lens 17, the second relay lens 16 and the first relay lens 15, and again passes through the two-dimensional scanning device 14.
Passes through the quarter-wave plate 13 and the polarizing beam splitter 12 through the optical path and travels toward the second imaging lens 18. The laser light L1 is condensed by the second imaging lens 18, passes through the optical aperture 19a having a pinhole, and
The light enters the light receiving element 19b. The first light receiving element 19b is composed of, for example, a photomultiplier or a photodiode. The first light receiving element 19b receives the incident laser light L1 by the light receiving element, converts the voltage with an output current corresponding to the amount of incident light through a current-voltage conversion circuit, and converts the signal. The signal is output to the first A / D conversion circuit 41 via a gain amplification circuit 46 (see FIG. 1) that performs amplification and gain control.

Next, luminance information obtained by the laser optical system 1 will be described. The light stop 19a is disposed at the focal position of the second imaging lens 18, and the pinhole of the light stop 19a is extremely small, so that the laser beam L1 focuses on the sample w. And the reflected light L1
Forms an image at the pinhole of the optical diaphragm 19a, and the amount of received light incident on the first light receiving element 19b is significantly increased. vice versa,
If the laser light L1 is not focused on the sample w, the reflected light L1 hardly passes through the pinhole of the optical stop 19a, so that the amount of light received by the first light receiving element 19b becomes extremely small.

FIG. 6 is a diagram showing the relationship between the brightness of the microscope and the resolving power. FIG. 6A is a diagram showing the relationship between brightness and resolution of a confocal microscope, and FIG. 6B is a diagram showing the relationship between brightness and resolution of a normal microscope. As can be seen from FIG. 6A, when the confocal optical system is in focus, most of the light passes through the pinhole, but when out of focus, the amount of light passing through the pinhole is It is a drastic decrease. On the other hand, in the ordinary microscope of FIG. 6B, the resolving power peaks at the focal position, but the brightness does not change much. Thus, when the confocal optical system is in focus, the brightness and the resolving power peak at the focal position. In other words, the maximum luminance and the in-focus position match, so that a bright image is obtained at the in-focus portion, while a dark image is obtained at other out-of-focus positions. As described above, the confocal microscope can obtain an image with an extremely shallow depth, and can obtain a bright image in which only the target layer is in focus, and a dark image can be obtained for the other layers. be able to. In addition, sample
When the image is moved in the axial direction and the maximum luminance signal is stored in each pixel of the image, the image is focused on the entire surface, and conversely, an image with an infinite depth of focus can be obtained. In addition, by storing height information of the focused Z-axis at each point of the scanning line, the surface shape can be measured.

Next, the white light optical system 2 will be described. The white light optical system 2 includes a white light (illumination light for color information) L
2 is used as a light source. On the optical axis of the white light source 20, a second collimating lens 21,
A first half mirror 22 and the objective lens 17 are provided.
The white light optical system 2 is disposed so that the optical axes of the two optical systems 1 and 2 match. Therefore, the white light L2 is focused on the same place as the scanning area of the laser light L1. Sample w
The white light (response light) L2 reflected by the light source passes through the objective lens 17, the first half mirror 22, and the second relay lens 16, is further reflected by the second half mirror 23, and is reflected by the color CCD ( An image is formed on the surface of the (second light receiving element) 24. That is, the color CCD 24 is connected to the light diaphragm 1
It is arranged at a position conjugate with or close to conjugate with 9a. The image picked up by the color CCD 24 is read out by the CCD drive circuit 43 as analog color image pickup information and output to the second A / D conversion circuit 42.

Next, the color video signal generating means 5 shown in FIG.
Will be described. The color video signal creating means 5 includes the first
By combining the luminance information from the light receiving element 19b and the color information from the color CCD 24, digital signals ro, go, and bo for color images are created. The color video signal creating means 5 includes first and second area circuits 5
1 and 52 and a luminance conversion circuit 53 and the like. As shown in FIG. 8, the first and second area circuits 51 and 52 each use a predetermined common part of the imaging area A1 of the laser optical system 1 and the imaging area A2 of the white light optical system 2 as an image area A0. Select and output a digital signal for the selected portion. That is, the first area circuit 51 of FIG.
For the image area A0, the luminance signal i is stored in the luminance memory Mi at a resolution corresponding to each pixel of the color CCD 24. On the other hand, the second area circuit 52 stores the red, green, and blue color intensity signals rm, gm, bm for each pixel in the video area A0 in the first color intensity memories Mr1, Mg1,
It is stored in Mb1. Note that the color intensity signal is a signal including luminance (intensity) of the three primary colors.

The luminance conversion circuit 53 uses the following equation (1),
According to (2) and (3), the converted color intensity signals ro, go, and bo are obtained by replacing the luminance information of the color intensity signals rm, gm, and bm for each pixel with the luminance information of the luminance signal i. The converted color intensity signals ro, go, and bo are stored in the second color intensity memories Mr2, Mg2, and Mb2. Ro = I · Rm / (Rm + Gm + Bm) (1) Go = I · Gm / (Rm + Gm + Bm) (2) Bo = I · Bm / (Rm + Gm + Bm) (3) where I: Luminance of luminance signal i Rm, Gm, Bm: luminance (intensity) of color intensity signals rm, gm, bm Ro, Go, Bo: luminance (intensity) of converted color intensity signals ro, go, bo Note that the first color intensity The memories Mr1, Mg1, Mb1 and the second color intensity memories Mr2, Mg2, Mb2 are color CCs.
D24 has a storage unit corresponding to the pixels in the above-described image area A0. The converted color intensity signals ro, go, and bo thus obtained are signals obtained by replacing the luminance information in the color imaging information from the color CCD 24 with the luminance information from the first light receiving element 19b. The converted color intensity signals ro, go, bo are stored in the second color intensity memories Mr2, M
g2, Mb2 and read out from the display controller 72.
Is output to the D / A conversion circuit 60 constituting the display controller 72. Further, the synchronizing signal a is added in the adder 61 constituting the display controller 72 to become an analog composite color video signal c. This composite color video signal c is displayed on a display device (monitor).
The image is output to 62 and an image of the sample w is displayed.

Next, how to use the present color confocal microscope will be described. The present color confocal microscope selects and uses one of three modes: an area search mode using a CCD camera, a black-and-white (achromatic) confocal image mode, and a color confocal image mode. The settings of these modes are set by operating the above-mentioned icons displayed on the display screen with the operation unit (for example, a mouse) 63.

When the area search mode is selected, the color video signal creating means 5 stops the laser drive circuit 44 shown in FIG. 5 and activates the CCD drive circuit 43 to cause the color CCD 24 to pick up an image. In this area search mode, the first color intensity memory Mr from the second area circuit 52 in FIG.
1, color intensity signals rm, g stored in Mg1, Mb1
m and bm are output to the D / A conversion circuit 60 as they are, and a normal enlarged image with a large depth of field is displayed on the monitor 62. Therefore, by moving the sample stage 30 in FIG. 5 in the X and Y directions, it is possible to search for an area to be imaged.

When the black-and-white confocal image mode is selected, the color video signal generating means 5 (FIG. 7) controls the laser driving circuit 44 of the laser optical system 1 and the two-dimensional scanning device 1.
4 and the like, and an image is taken by the laser optical system 1.
In the monochrome confocal image mode, the first area circuit 5 shown in FIG.
The luminance signal i stored in the luminance memory Mi from 1 is output to the D / A conversion circuit 60 as it is, and a black-and-white (achromatic) enlarged image with high resolution is displayed on the monitor 62.

Further, when the color confocal image mode is selected, the laser driving circuit 44 and the CCD driving circuit 43
Are driven alternately. That is, when the color confocal image mode is selected, the process proceeds to step S92 in step S91 of FIG. 9, and after one screen is scanned by the laser beam L1, the process proceeds to step S93. Step S93
5, the laser drive circuit 44 of FIG. 5 stops, and the laser beam L1 is no longer emitted from the laser 10. In this state, the process proceeds to step S94 in FIG. The color intensity signal r of FIG. 7 obtained in step S94
For m, gm, and bm, the luminance information included in this signal is replaced with the luminance information of the luminance signal i obtained in step S92, and becomes the converted color intensity signals ro, go, and bo. The converted color intensity signals ro, go, and bo are stored in the second color intensity memories Mr2, Mg2, and Mb2, respectively, and then stored in D /
The color enlarged image is output to the A conversion circuit 60 and displayed on the display screen 62. Note that, after step S94 in FIG. 9, the process proceeds to step S95, in which the scanning with the laser light L1 and the accumulation and readout of charges by the CCD drive circuit 43 are repeated. Since the color confocal image obtained in this way has a low resolution for color information, it has a slightly lower resolution than a conventional color laser microscope using laser light of three primary colors, but has a higher resolution than a normal enlarged image. In this way, images with sufficiently high utility value can be obtained.

Also, the white light source 20 and the color CCD shown in FIG.
Since color information is obtained by the white light optical system 2 using the light source 24, the optical system has a significantly simpler structure than a conventional color laser microscope using laser light of three primary colors, so that the cost and size of the microscope can be reduced. I can figure it out. When the image is picked up by the color CCD 24 shown in FIG. 5, the laser driving circuit 44 is stopped so that the laser light L1 does not enter the color CCD 24. And an image of a color close to the actual color of the sample w is obtained. As a means for preventing the laser light L1 from being incident on the color CCD 24, various methods such as using a shutter for shielding the laser light L1 or setting the scanning range of the laser light L1 outside the imaging area of the color CCD 24 are available. Can be adopted.

Although the color CCD 24 is used as the second light receiving element of the second optical system 2 in FIG. 5, another light receiving element may be used. For example, the reflected light L2 is separated into three primary colors by using a dichroic mirror, and the reflected light of these three primary colors is converted into three primary colors.
The light may be incident on two two-dimensional CCDs for monochrome images. Although the optical system is different, a one-dimensional scanning device that uses a color line CCD as the second light receiving element and one-dimensionally scans white response light may be provided. Further, as the second light receiving elements, three line CCDs (R, G, B) for monochrome images are used.
) Can be used, and three point light receiving elements (for R, G, B) can also be used. In these cases, the scanning device that scans white response light can also be used as the scanning device that scans laser light L1. Further, as the second light receiving element, in addition to the color CCD, another solid-state imaging element such as a MOS type, a television camera in which a plurality of imaging tubes are combined, or the like can be used. FIG.
In the first embodiment, the first light receiving element 19b and the color CCD 24 receive the reflected light of the laser light L1 and the white light L2, respectively. However, the transmitted light transmitted through the sample w and the fluorescence substituted for the reflected light are received. It may be something. Although the colors are separated into the three primary colors of light here, they may be separated into complementary colors (yellow, cyan, green). Further, a color difference signal may be used as the color information. As described above, according to the present microscope according to the prior invention, color information is obtained by the second optical system using illumination light for color information different from laser light, so that the conventional color laser microscope using laser light of three primary colors is used. Since the optical system has a remarkably simple structure, the cost can be reduced and the size can be reduced.

In the scanning in the above description, the laser beam or the line laser beam is scanned while the sample is stationary, but the sample may be moved without scanning the laser beam or the line laser beam. The sample may be moved in the Y direction (a direction orthogonal to the X direction) by scanning the light in the X direction. The signal for a color image is a composite signal including a video signal composed of the intensities of the three primary colors of light (red, green, and blue), a signal composed of a luminance signal and a color difference signal, and a horizontal synchronization signal and a color burst signal. It refers to a signal capable of displaying a color image as it is or after being processed, such as a color image signal. Also, the luminance information is
Refers to information about luminance that does not include color.
For example, it refers to information on the balance of color intensities, such as a color difference signal.

[0018]

In such a confocal microscope, if the amplification factor (gain) of the amplifier circuit for amplifying the output value of the first light receiving element is small, the input voltage to the first A / D conversion circuit is small. That is, the measurement is incomplete, and the amplification factor must be set again and the measurement must be performed again. If the amplification factor is too large, the input voltage to the first A / D conversion circuit is saturated, Both bright and dark luminance parts are displayed brightly, making it impossible to distinguish the unevenness of the surface of the object (work), and it is necessary to set the amplification factor again and perform the measurement again. I got up and my work efficiency dropped. In addition, selecting the amplification factor of the amplifier circuit to the optimum value itself is time-consuming because it is difficult.

Also, there are published publications describing automatic gain adjustment and automatic offset voltage adjustment. this is,
A shutter that can be shut off on the optical path between the light source and the work is provided, and all the image data of a plurality of images having different distances in the Z direction are taken into the memory while the light is shut off, or the image data in the Z direction is opened when the light is released. Multiple image data with different distances were all loaded into the memory, and the average value, maximum value, minimum value, etc. were calculated using these data to determine the offset value and the optimal gain. . The gain adjustment and the offset adjustment by this method require a plurality of image data and require a large amount of memory, resulting in high cost.

An object of the present invention is to solve the above-mentioned drawbacks. A confocal microscope that does not require a large amount of memory and that can adjust gain only in a soft level in a short time and reliably. To provide an automatic adjustment method for

[0021]

In order to achieve the above object, the invention of a method for automatically adjusting a confocal microscope according to claim 1 of the present invention is to focus light on a sample by an objective lens and to reduce response light by the light. In a confocal microscope in which light is received by a light receiving element, the gain of the light receiving element is automatically adjusted by a search method from information on one screen.

According to a second aspect of the present invention, there is provided a variable gain amplifying circuit having a variable gain for amplifying a light receiving signal of a light receiving element, and a signal amplified by the variable gain amplifying circuit, the signal being amplified by an A.
A / D converter circuit for performing A / D conversion, a frame memory for storing an output value of the A / D converter circuit, a gain memory for storing a gain of the variable gain amplifier circuit, and the amplification based on the gain. D / for changing the gain of the circuit
An A converter, a set target value memory for storing a set target value which is a set target for an output value stored in the frame memory or a value obtained from the output value, and an output value stored in the frame memory A CPU for calculating a gain of the variable gain amplifier circuit for calculating a value obtained from the value to the set target value, and the CPU is stored in the frame memory. The variable gain amplifier circuit calculates the gain of the variable gain amplifier circuit using a search method for setting an output value or a value obtained from the output target value to the set target value, and the variable gain amplifier circuit set to the gain obtained by the calculation. Then, the output value stored in the frame memory is updated with the output value obtained by amplifying the light receiving signal of the light receiving element again and performing A / D conversion by the A / D converter circuit. The output value stored in the frame memory or a value obtained from the value is compared with the set target value, and if the result of the comparison does not match, the calculation and comparison of the CPU are repeated to approach the set target value. The automatic adjustment of the gain is automatically obtained.

Further, the invention according to claim 3 is the same as the invention according to claim 2.
In the automatic adjustment method for a confocal microscope described above, the search method first uses a half of a sum of a minimum adjustment value and a maximum adjustment value of the gain variable D / A converter circuit as a gain adjustment value. The output of the light receiving element is stored in the frame memory from the A / D converter circuit, and the maximum measured value of the stored value is compared with the target value. If the value is smaller than the value, the gain of the gain amplifier circuit is changed using a new gain adjustment value that is added to the gain adjustment value with 1/2 of the gain adjustment value as an increase / decrease width. Is stored in the frame memory from the A / D converter circuit, and the maximum measured value of the stored value is compared with the target value again. Also, the maximum measured value of the calculation result is smaller than the target value. When big, gay The output of the first light receiving element is calculated using the new gain adjustment value subtracted from the gain adjustment value with 1/2 of the adjustment value as an increase / decrease width, and the maximum measurement value of the calculation result is again compared with the target value. The result of the comparison between the maximum measurement value of the calculation result and the target value in the or the above, when the maximum measurement value of the calculation result is smaller than the target value, returns to the above, and the maximum measurement value of the calculation result is the If the target value is larger than the target value, return to the above, repeat the above, and when the maximum measured value of the calculation result matches the target value or when the gain adjustment value falls below the predetermined gain value, terminate the search. It is characterized in that an adjustment value is used.

The invention described in claim 4 is the same as the claim 2.
In the automatic adjustment method for a confocal microscope described above, the search method first changes the gain of the gain amplification circuit with a predetermined gain adjustment value, and outputs the output of the first light receiving element at this time to the A / D converter. Stored in the frame memory from the circuit,
Comparing the maximum measured value of the stored value with the target value; and, as a result of the comparison, when the maximum measured value of the operation result is smaller than the target value, increasing or decreasing a predetermined unit gain to the predetermined gain adjustment value. The gain of the gain amplifier circuit is changed using the new gain adjustment value added as the width, and the output of the first light receiving element at this time is stored in the frame memory from the A / D converter circuit, and the stored data is stored. Is compared again with the target value, and if the result of the comparison is still smaller than the target value, a new gain adjustment value obtained by further adding a predetermined unit gain to the new gain adjustment value as an increase / decrease range Is used to change the gain of the gain amplifier circuit, the output of the first light receiving element at this time is stored in the frame memory from the A / D converter circuit, and the maximum measured value of the stored value and the Target value Is compared again, and the above is repeated.When the maximum measured value of the calculation result matches or exceeds the target value, the search is terminated, and the gain adjustment value at the time of is used. When the maximum measured value of the calculation result is larger than the target value, the gain of the gain amplifying circuit is changed using a new gain adjustment value obtained by subtracting a predetermined unit gain from the predetermined gain adjustment value as an increase / decrease width. The output of the first light receiving element at this time is stored in the frame memory from the A / D converter circuit, and the maximum measured value of the stored value is again compared with the target value. When the gain is still larger than the target value, the gain of the gain amplifier circuit is changed using a new gain adjustment value obtained by further subtracting a predetermined unit gain from the new gain adjustment value as an increase / decrease range, and the first gain at this time is changed. The output of the optical element is stored in the frame memory from the A / D converter circuit, the maximum measured value of the stored value is compared with the target value again, and the above is repeated. The search is terminated when the value matches or falls below the target value, and the gain adjustment value at the time is used. According to a fifth aspect of the present invention, there is provided a method for automatically adjusting a confocal microscope, comprising a search method according to a third aspect and a search method according to a fourth aspect.

Further, according to the invention of a confocal microscope according to claim 6, a variable gain amplifier circuit having a variable gain for amplifying a light receiving signal of a light receiving element, and A / D converting the signal amplified by the variable gain amplifier circuit. A / D converter circuit, a frame memory for storing an output value of the A / D converter circuit, a gain memory for storing a gain of the variable gain amplifier circuit, and a gain of the amplifier circuit based on the gain. A D / A converter for varying the output value, a set target value memory for storing a set target value which is a set target for an output value stored in the frame memory or a value obtained from the value, and the frame memory CPU for calculating the gain of the variable gain amplifying circuit for calculating the output value stored in the memory or the value obtained from the value to the set target value
A confocal microscope, wherein the CPU searches for a gain of the variable gain amplifier circuit for setting the output value stored in the frame memory or a value obtained from the value to the set target value. Calculated using the variable gain amplifier circuit set to the gain obtained by the calculation,
The output value stored in the frame memory or a value obtained from the output value obtained by amplifying the light receiving signal of the light receiving element again and performing A / D conversion by the A / D converter circuit is referred to as the set target value. If the result of the comparison does not match, the calculation and the comparison of the CPU are repeated, and the adjustment of the gain is automatically obtained so as to approach the set target value. According to a seventh aspect of the present invention, when the confocal microscope according to the sixth aspect includes a display device for displaying the contents of the frame memory, and when a dark portion of a display image on the display device is difficult to see, It is characterized in that an offset value can be added to the entire display image data in the memory.

According to the above configuration, in the confocal microscope, the gain can be automatically adjusted in a short time in a short time with only the soft level from the information of one screen, and the work efficiency can be improved as in the conventional technique. Does not decrease.

[0027]

FIG. 1 is a block diagram showing an embodiment of the present invention.
This will be described with reference to FIG. FIG. 1 shows a block diagram of a confocal microscope of the present invention. In FIG. 1, reference numeral 80 denotes a control unit, and the control unit 80 includes a CPU 81, a RAM 8
2, a ROM 83, a frame memory 84, an interface 85, a display controller 86, a gain memory 87, and a target value memory 88. Although the frame memory 84, the gain memory 87, and the target value memory 88 are illustrated independently in FIG. 1 for easy-to-understand functions, they may actually be provided in the RAM 82. The control unit 80 includes the first A / D converter 41 and the second A / D converter 42 described above with reference to FIG. 5, and an operation unit (for example, a mouse, a keyboard, a trackball, etc.) 6.
3 is connected as an input, and similarly, the laser light drive circuit 44, the two-dimensional scanning device 14, the stage control circuit 40, and the display device 62 such as a CRT, an LCD, etc.
Are connected as outputs. The first A / D converter 41 adds the offset voltage output from the D / A conversion circuit 47 and the voltage obtained by current-to-voltage conversion of the output signal from the first light receiving element 19b by the addition circuit 45. Is further input by a variable gain amplifier circuit 46 with a predetermined gain. On the other hand, the output signal from the second light receiving element 43 is supplied to the second A / D converter 42 by the amplifier circuit 431.
Is amplified and input. The CPU 81 controls the RAM 82 and the frame memory 84 and performs data processing according to the arithmetic processing procedure stored in the ROM 83. The RAM 82 stores data to be processed by the CPU 81 and processed data. The ROM 83 is a CPU 8
1 is stored. The frame memory 84
The output values from the first A / D conversion circuit and the second A / D conversion circuit transmitted and received from the interface 85 are stored. The display controller 86 controls the display device 62 to display the bitmap data in the frame memory 84. The gain memory 87 is a memory for storing the gain of the variable gain amplifier circuit 46 described above. The target value memory 88 is a memory for storing a set target value which is a set target for the output value stored in the frame memory 84 or a value obtained from the output value. The display device 62 displays the bitmap data in the frame memory 84. Operation unit 63
Is for writing various programs and various parameters.
Used when reading, creating, changing, monitoring, saving, executing, etc.

The gain adjustment is performed when the laser 10 (FIG. 5) is turned on and the focus of the objective lens 17 just coincides with the work w, that is, when the maximum light amount is obtained. When the signal enters the element 19b, the maximum value of the signal is set to the value of the target value memory of the input of the A / D conversion circuit 41. For example, when the value of the target value memory is set to 255, if the input of the A / D conversion circuit 41 is 8 bits, the maximum value is 255. Therefore, the maximum value of the signal is set to the input value of the A / D conversion circuit. Adjustment to bring the maximum value level to 255. For automatic gain adjustment, the CPU 81 adjusts the variable gain amplifier circuit 46 so that the output value stored in the frame memory 84 or a value obtained from the value matches the set target value stored in the target value memory 88. Is calculated by an optimum gain search method described later. Specifically, the variable gain amplifying circuit 46 is set via the variable gain D / A with the gain obtained by the calculation by the search method for the optimum gain, and the light receiving signal of the light receiving element 19b is again amplified with this gain ( 46) in FIG. 1 and the A /
The D / A converter circuit 41 compares whether the output value obtained by the A / D conversion matches the set target value, and if not, repeats the calculation and comparison of the CPU (81 in FIG. 1) to finally set the set target value. It approaches the value.

According to the present invention, the automatic gain adjustment is performed quickly and accurately by using the binary search method and / or the successively decreasing method. The algorithm is shown in the flowcharts of FIGS. 2 and 3, respectively. is there. FIG. 2 is a flowchart of the automatic gain adjustment by the binary search method.
FIG. 3 shows a flowchart of the automatic gain adjustment by the successively decreasing method. In FIG. 2, when this flow starts, the power is turned on in step S21, and the laser is turned on in step S22. In step S23, the target value, the gain adjustment value, and the increase / decrease range are initialized.
First, the target value is set to the maximum value that can be set, the gain adjustment value is set to ×× (minimum adjustment value + maximum adjustment value), and the increase / decrease range is set to the gain adjustment value. In step S24, laser screen measurement is performed using this gain adjustment value.
At 25, the next change width is set to の of the previous change width. In step S26, it is compared whether the increase or decrease is smaller than a desired value (for example, 1 of the minimum bit). If the increase or decrease is not smaller than the desired value, the process proceeds to step S28 to determine whether the maximum measured value of the laser screen exceeds the target value. If the answer is NO, the process proceeds to step S291, and a value obtained by adding the increase / decrease width to the gain adjustment value is set as a new gain adjustment value. If YES, the process proceeds to step S292, and a value obtained by subtracting the increase / decrease width from the gain adjustment value is set as a new gain adjustment value. Then, the process returns to step S24, and the laser screen measurement is performed using the gain adjustment value. If the increase / decrease width is smaller than the desired value in step S28, no further search can be performed, and the search is terminated (step S27), and the gain adjustment value at that time is used in subsequent measurements.

When the distance between the objective lens and the work is fixed in a state where the focus is almost in focus, the optimum gain can be adjusted by executing the binary search method shown in FIG. in this way,
According to the bisection search, the target value can be reached with a small number of searches. That is, assuming that the set range is n stages, the target value can be reached by [(log ₂ n) -1] searches at most. For example, if the setting range n of the A / D conversion circuit is 256 steps, (lo
g ₂ n) and _{-1 = log 2 256-1 = log 2} 2 8 -1 = 8-1 = 7, it is possible to reach the target value seven times of the search at the highest.

FIG. 3 shows a flow chart of the automatic gain adjustment by the progressive reduction method. In FIG. 3, when this flow starts, the power is turned on in step S31,
In step S32, the laser is turned on. After turning on the laser, the target value, the gain adjustment value, and the increase / decrease range are initialized in step S33. First, the target value is set to a settable maximum value, the gain adjustment value is set to a predetermined gain adjustment value, and the increase / decrease range is set to an arbitrary predetermined gain value. In step S34, laser screen measurement is performed using the current gain adjustment value. In step S35, a comparison is made as to whether the maximum measured value of the laser screen exceeds the target value. If the comparison result is NO, the process proceeds to step S36. Step S36
Then, a value obtained by adding the increase / decrease width to the gain adjustment value is set as a new gain adjustment value (step S36). Then, it is determined whether the new gain adjustment value is equal to or greater than the maximum gain value (step S361). As a result of the comparison, if the new gain adjustment value is equal to or greater than the maximum gain value, the adjustment is terminated because there is no way to increase the gain anymore (proceed to step S38), and the maximum gain value is changed to a variable gain amplifier circuit thereafter. Is adopted as the gain of On the other hand, if the result of the comparison indicates that the new gain adjustment value is lower than the maximum gain value, the laser screen measurement is performed again with the new gain adjustment value (step S).
362). In step S363, whether the maximum measured value on the laser screen exceeds the target value is compared again, and the comparison result is N
If O (ie, if not exceeded), step S
Proceed to 36 and repeat the same procedure. On the other hand, if the maximum measured value on the laser screen exceeds the target value in step S363, the search ends (step S38). In this case, the gain value to be adopted as the gain of the variable gain amplifier circuit is based on the margin of the target value, whether the gain adjustment value is closer to the target value, or the like. It is only necessary to determine which of the gain adjustment values immediately before the loop is to be used. Alternatively, the search may be started by the bisection method from here until the maximum measured value on the laser screen matches the target value, and a more accurate gain adjustment value may be determined. On the other hand, if the comparison result in step S35 is YES (that is, if the maximum measured value of the laser screen exceeds the target value), the process proceeds to step S37. In step S37, a value obtained by subtracting the increase / decrease width from the gain adjustment value is set as a new gain adjustment value (step S37). Then, it is determined whether the new gain adjustment value is equal to or less than the minimum gain value (step S37).
1). As a result of the comparison, if the new gain adjustment value is equal to or smaller than the minimum gain value, the adjustment is terminated because there is no way to lower the gain anymore (proceed to step S38), and the minimum gain value is changed to a variable gain amplifier circuit thereafter. Is adopted as the gain of On the other hand, if the result of the comparison indicates that the new gain adjustment value exceeds the minimum gain value, the laser screen measurement is performed again with the new gain adjustment value (step S372). Step S
At 373, the comparison is again performed to determine whether the maximum measured value on the laser screen exceeds the target value. If the comparison result is YES (that is, if the maximum value is exceeded), the process proceeds to step S36, and the same procedure is repeated thereafter. On the other hand, if the maximum measured value of the laser screen does not exceed the target value in step S363, the search ends (step S38). In this case, the gain value to be adopted as the gain of the variable gain amplifier circuit is based on the margin of the target value, whether the gain adjustment value is closer to the target value, or the like. It is only necessary to determine which of the gain adjustment values immediately before the loop is to be used. Alternatively, the search may be started by the bisection method from here until the maximum measured value on the laser screen matches the target value, and a more accurate gain adjustment value may be determined.

While the binary search method of FIG. 2 is convenient to use when the focal point of the objective lens is fixed to exactly match the workpiece, the search method of FIG. Even if the focus does not match the work, it can be adjusted. That is, when the focus of the objective lens does not coincide with the work, the input signal of the light receiving circuit is A / D
The gain of the conversion circuit is set to the maximum level, and thereafter, the distance between the objective lens and the work is changed in a direction in which the input signal of the light receiving circuit increases (that is, in a direction in which the focus of the objective lens approaches the work). Since the input signal of the circuit increases, the gain of the A / D conversion circuit is lowered each time, the input signal at that time is set to a target set value, and this is repeated. Even if the number of signals increases, the maximum level is updated each time. Therefore, there is no need to store a plurality of pieces of image data in a memory as in the conventional invention.

According to the head-sequential decreasing search, when the set range is n stages, the number of searches to reach the target value is the highest, n + 1 times, and the expected value is [(n + 1) / 2] +1 times the lowest. One time. For example, when the setting range n of the A / D conversion circuit is 256 steps, the expected value is 257 times at the maximum, and the expected value is 129 times at the minimum, and 1 time.

The two methods of the binary search and the successively decreasing method are described.
It is better to combine them. For example, when the prediction of the setting value such as the slice screen measurement is not known at all, the setting value is determined by the dichotomous search. Conversely, when the prediction of the setting value such as the super depth screen measurement can be obtained to some extent, It is better to check the set value from the prediction point. Also, in the case of the ultra-depth screen measurement, when the set value cannot be predicted, a certain degree of predicted value is set by the dichotomous search, the search is performed monotonously and coarsely in order from the predicted value, and finally the dichotomous search is performed again. You can reach the target value quickly.

The work displayed on the display screen may be dark and difficult to see due to the deepness of the recess, but the conventional offset voltage adjustment is performed, for example, in the actual open square 7-2.
As described in Japanese Patent No. 0382, only subtraction is performed. As a result, as a result of subtracting the offset voltage, the portion where the light receiving signal voltage is low becomes darker and hard to see. Conversely, if the gain is increased to make the portion bright, the light receiving signal voltage is high, the bright portion is saturated, and the unevenness on the surface of the bright portion is crushed.

FIG. 4 shows another embodiment of the present invention which simply solves the above drawbacks. That is, the work displayed on the screen may be dark and difficult to see. However, according to the present invention, a portion where the light reception signal voltage is low becomes bright and is easily seen, and a bright portion has unevenness on the surface. No more. FIG. 4 is a diagram illustrating this state. In FIG. 4A, since there is a portion where the light receiving signal voltage is low,
FIG. 4 shows a state in which the part of the work is dark and difficult to see.
FIG. 4B shows a solution of a conventional example in which a dark and hardly visible portion in FIG. 4A is made easier to see, and FIG.
2 shows a solution according to the invention for making dark and hard-to-see places easier to see. In FIG. 4B of the conventional example, FIG.
The dark spot a3 in (A), which is hard to see, has a light receiving signal voltage increased as shown by b3 due to the integration of a predetermined gain, so that it is certainly easy to see. However, conversely, the light receiving signal voltage is saturated at the points a1 and a2, which were clearly distinguished in FIG. 4A, as indicated by b1 and b2 due to the product of the predetermined gain, and the unevenness on the bright surface is obtained. Has become unrecognizable. However, in FIG. 4C according to the present invention, instead of obtaining a product of a predetermined gain as in FIG. 4B of the conventional example, the offset voltage obtained by the above-described method is added to the whole. I have. As a result, the voltage at the portion where the light receiving signal voltage is low is raised (see c3), and the portion becomes bright and easily visible. Also, originally bright portions such as a1 and a2 in FIG. 4A do not perform gain integration as in the prior art, so that saturation such as b1 and b2 does not occur, and concavities and convexities are crushed. (Note the difference in waveform between b1 and c1 and between b2 and c3).

[0037]

As described above, according to the present invention, in a confocal microscope in which light is condensed on a sample by an objective lens and response light by the light is received by a light receiving element, the gain of the light receiving element is improved. Since the adjustment is automatically performed from the information of one screen by the search method, the gain can be automatically adjusted in only a soft level in a short period of time and reliably. Becomes unnecessary, and a decrease in work efficiency can be prevented.

[Brief description of the drawings]

FIG. 1 is a block diagram of a confocal microscope of the present invention.

FIG. 2 is a flowchart of automatic gain adjustment by a binary search method.

FIG. 3 is a flowchart of an automatic gain adjustment by a successively decreasing method.

FIG. 4 is a diagram illustrating a state of a light receiving signal voltage of a work having a dark and hardly visible portion.

FIG. 5 is a configuration diagram schematically showing a color confocal microscope which is a basis of the present invention.

6A is a diagram showing the relationship between brightness and resolution of a confocal microscope, and FIG. 6B is a diagram showing the relationship between brightness and resolution of a normal microscope.

FIG. 7 is a block diagram of a color video signal generating unit of the confocal microscope.

FIG. 8 is a plan view showing an imaging area of the color confocal microscope.

FIG. 9: Laser drive circuit and CCD of color confocal microscope
6 is a flowchart illustrating a procedure in which a driving circuit and a driving circuit are alternately driven to obtain a composite screen.

[Explanation of symbols]

Reference Signs List 1 laser optical system (first optical system) 10 laser emitting laser light L1 11 first collimating lens 12 polarizing beam splitter 13 quarter-wave plate 14 two-dimensional scanning device 15 first relay lens 16 second relay lens 17 objective Lens 18 Second imaging lens 19a Optical aperture unit having a pinhole 19b First light receiving element 2 White light optical system (second optical system) 20 White light source that emits white light 21 Second collimating lens 22 First half mirror 23 Second half mirror 24 Color CCD (second light receiving element) 30 Sample stage 41 First A / D conversion circuit 42 Second A / D conversion circuit 43 CCD drive circuit 431 Amplification circuit 46 Variable gain amplification circuit 47 D / A conversion circuit 5 Color Video signal generating means 51 First area circuit 52 Second area circuit 53 Brightness conversion circuit 61 Addition Device 62 display device (monitor) 72 display controller 80 control unit 81 CPU 82 RAM 83 ROM 84 frame memory 85 interface 86 display controller 87 gain memory 88 target value memory L1 reflected light L2 white light A0 common image area A1 laser optical system Imaging area A2 Imaging area of white light optical system c Signal for color image (composite color image signal) i Luminance signal w Work (sample) ro, go, bo Digital signal for color image (converted color intensity signal) rm, gm , Bm Red, green, blue color intensity signals Mi Luminance memory Mr1, Mg1, Mb1 First color intensity memory Mr2, Mg2, Mb2 Second color intensity memory

Claims (7)

[Claims] 1. A confocal microscope in which light is condensed on a sample by an objective lens and response light by the light is received by a light receiving element, wherein a gain adjustment of the light receiving element is automatically performed by a search technique from information on one screen. A method for automatically adjusting a confocal microscope. 2. A variable gain amplifying circuit having a variable gain for amplifying a light receiving signal of a light receiving element, and an A / D converter for amplifying the signal amplified by the variable gain amplifying circuit.
An A / D converter circuit for converting; a frame memory for storing an output value of the A / D converter circuit; a gain memory for storing a gain of the variable gain amplifier circuit; A D / A converter for changing a gain; a set target value memory for storing a set target value which is a set target for an output value stored in the frame memory or a value obtained from the value; A CPU for calculating a gain of the variable gain amplifier circuit for calculating the output value stored in the memory or a value obtained from the value to the set target value;
A confocal microscope comprising: a method of searching for a gain of the variable gain amplifying circuit for setting an output value stored in the frame memory or a value obtained from the value to the set target value. The variable gain amplifying circuit set to the gain obtained by the calculation amplifies the light receiving signal of the light receiving element again, and obtains the output value obtained by A / D conversion by the A / D converter circuit. The output value stored in the frame memory is updated, and the output value stored in the frame memory or a value obtained from the value is compared with the set target value. An automatic adjustment method for a confocal microscope, wherein calculation and comparison are repeated to automatically obtain a gain adjustment so as to approach the set target value. 3. The automatic adjustment method for a confocal microscope according to claim 2, wherein the search method firstly uses 1/1/2 of the sum of a minimum adjustment value and a maximum adjustment value of the gain variable D / A converter circuit. 2 as a gain adjustment value, storing the output of the first light receiving element from the A / D converter circuit in the frame memory, comparing the maximum measured value of the stored value with the target value, When the maximum measured value of the result is smaller than the target value, the gain of the gain amplifying circuit is changed by using a new gain adjustment value that is added to the gain adjustment value with １ ／ of the gain adjustment value as an increase / decrease range. The output of the first light receiving element at this time is stored in the frame memory from the A / D converter circuit, and the maximum measured value of the stored value is again compared with the target value. Maximum measured value is greater than the target value When the threshold value is larger, the output of the first light receiving element is calculated using the new gain adjustment value obtained by subtracting the gain adjustment value from 1/2 of the gain adjustment value as an increase / decrease range. Comparing again with the target value, as a result of comparing the maximum measured value of the operation result or the target value with the target value, when the maximum measured value of the operation result is smaller than the target value, returning to the above; When the maximum measured value is larger than the target value, the process returns to the above, and the above operation is repeated. When the maximum measured value of the calculation result matches the target value or the gain adjustment value falls below the predetermined gain value, the search is terminated. Automatic adjustment method of confocal microscope. 4. The automatic adjustment method for a confocal microscope according to claim 2, wherein the search method first changes the gain of the gain amplifier circuit with a predetermined gain adjustment value, and then changes the gain of the first light receiving element. Storing the output from the A / D converter circuit in the frame memory;
Comparing the maximum measured value of the stored value with the target value; and, as a result of the comparison, when the maximum measured value of the operation result is smaller than the target value, increasing or decreasing a predetermined unit gain to the predetermined gain adjustment value. A new gain adjustment value added as a width is calculated, and if the new gain adjustment value is equal to or larger than the maximum gain value, the search is terminated; otherwise, the gain of the gain amplifier circuit is calculated using the new gain adjustment value. The output of the first light receiving element at this time is stored in the frame memory from the A / D converter circuit, and the maximum measured value of the stored value is again compared with the target value. When the result is still smaller than the target value, a new gain adjustment value is further calculated by adding a predetermined unit gain to the new gain adjustment value as an increase / decrease range, and when the new gain adjustment value is equal to or more than the maximum gain value, Finish the search Otherwise, the gain of the gain amplifier circuit is changed using the new gain adjustment value, and the output of the first light receiving element at this time is stored in the frame memory from the A / D converter circuit, Comparing the maximum measured value of the stored value with the target value again, repeating the above, and terminating the search when the maximum measured value of the calculation result matches or exceeds the target value; As a result, when the maximum measured value of the calculation result is larger than the target value, a new gain adjustment value is calculated by subtracting a predetermined unit gain from the predetermined gain adjustment value as an increase / decrease range, and the new gain adjustment value is calculated as the gain. If the value is equal to or smaller than the minimum value, the search is terminated; otherwise, the gain of the gain amplifier circuit is changed using the new gain adjustment value, and the output of the first light receiving element at this time is subjected to the A / D conversion. From the circuit Stored in the frame memory, again compares the maximum measured value of the stored value with the target value, and when the result of the comparison is still larger than the target value, increases or decreases a predetermined unit gain from the new gain adjustment value. A new gain adjustment value that is further subtracted as a width is calculated, and when the new gain adjustment value is equal to or smaller than the minimum gain value, the search is terminated. Otherwise, the new gain adjustment value is used to calculate the gain amplification circuit. The gain is changed, and the output of the first light receiving element at this time is stored in the frame memory from the A / D converter circuit, and the maximum measured value of the stored value is again compared with the target value. A method for automatically adjusting a confocal microscope, wherein the search is terminated when the maximum measured value of the calculation result coincides with or falls below the target value. 5. The search method according to claim 3, and claim 4.
3. The method for automatically adjusting a confocal microscope according to claim 1, further comprising the search method described in claim 1. 6. A variable gain amplifier circuit having a variable gain for amplifying a light receiving signal of a light receiving element, an A / D converter circuit for A / D converting a signal amplified by the variable gain amplifying circuit, and an A / D converter. A frame memory for storing an output value of the D converter circuit, a gain memory for storing a gain of the variable gain amplifier circuit, a D / A converter for changing a gain of the amplifier circuit based on the gain, A setting target value memory for storing a setting target value serving as a setting target for an output value stored in the frame memory or a value obtained from the value,
A CPU for calculating a gain of the variable gain amplifying circuit for calculating the output value stored in the frame memory or a value obtained from the value as the set target value;
A confocal microscope comprising: a method of searching for a gain of the variable gain amplifying circuit for setting an output value stored in the frame memory or a value obtained from the value to the set target value. The variable gain amplifying circuit set to the gain obtained by the calculation amplifies the light receiving signal of the light receiving element again, and obtains the output value obtained by A / D conversion by the A / D converter circuit. The output value stored in the frame memory is updated, and the output value stored in the frame memory or a value obtained from the value is compared with the set target value. A confocal microscope wherein a calculation and a comparison are repeated to automatically obtain a gain adjustment so as to approach the set target value. 7. The confocal microscope includes a display device for displaying the contents of the frame memory, and when a dark portion of the display image on the display device is difficult to see, an offset is applied to the entire display image data in the frame memory. 7. The confocal microscope according to claim 6, wherein values can be added.