JP2003140050A - Scanning confocal microscope - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a scanning confocal microscope by which three-dimensional images of good accuracy can be obtained and the color information of a sample surface can be reproduced. SOLUTION: This scanning confocal microscope has selecting means (21) for selecting the luminance signal of an optimum wavelength band from a plurality of the luminance signals varying in the wavelength bands of the light outputted after photoelectric conversion by imaging means and image building means (20) for building the three-dimensional images or the images of a deep depth of focus by utilizing the luminance signal of the optimum wavelength band.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a scanning confocal microscope utilizing the confocal effect.

[0002]

2. Description of the Related Art Generally, two types of scanning confocal microscopes are well known: a disk scanning type and a laser scanning type. Among them, the disc scanning confocal microscope has a high lateral resolution as compared with an ordinary microscope, and also has a very high sectioning effect in the optical axis direction of the sample (hereinafter, referred to as “Z direction”). It has features. And
By combining this feature with an image processing device, it is possible to construct a sample as a three-dimensional image.

FIG. 11 is a diagram showing the structure of a conventional disk scanning confocal microscope.

Illumination light emitted from the light source 1 enters a half mirror 3 through a collimator lens 2, and the light reflected here illuminates a rotating disk 4. As the rotating disk 4, a Nipkow disk in which a plurality of pinholes are spirally provided, or a disk in which a slit pattern is formed is used. Here, it is assumed that a Nipkow disk is used, and this rotating disk 4
Is attached to the rotating shaft 5a of the motor 5 and rotates at a predetermined rotation speed. Therefore, the illumination light applied to the rotary disc 4 passes through the plurality of pinholes formed in the rotary disc 4, and is imaged on the sample 7 by the objective lens 6.

The reflected light from the sample 7 passes through the objective lens 6 and the pinhole of the rotating disk 4 again, passes through the half mirror 3, and is focused on the image pickup section 9 by the condenser lens 8. The image capturing unit 9 captures the reflected light from the sample 7 and outputs the brightness signal to the computer 10.

The computer 10 fetches and stores the luminance signal output from the image pickup section 9, performs image processing to obtain desired image data and displays it on the monitor 11, and outputs a drive signal to the Z drive section 12. Then, a part or the whole of the optical system and the sample are relatively moved in the optical axis direction to change the focus position of the sample. Although only the objective lens 6 is moved here, the stage on which the sample 7 is loaded may be moved in the optical axis direction. Then, the in-focus position, which is position information in the Z direction, is also recorded in the computer 10 in association with the above-mentioned image data.

In the disk scanning confocal microscope thus constructed, the amount of light incident on the image pickup section 9 changes as the objective lens 6 moves, and when the surface of the sample comes into focus, the incident light amount ( Brightness) is maximum. Since each pixel of the image pickup device provided in the image pickup unit 9 outputs a luminance signal corresponding to the amount of light from each position of the sample 7, the sample 7 is obtained by obtaining the Z direction position showing the maximum luminance for each pixel.
Can be obtained, and by constructing the image only with the maximum luminance value of each pixel, it is possible to construct an image with a deep focal depth in which the entire surface of the sample 7 is in focus.

[0008]

In order for the image thus obtained to be clear with high accuracy, a curve showing the relationship between the luminance value and the position in the Z direction (hereinafter referred to as "IZ curve"). ) Is required to have steep characteristics.

FIG. 12 is a diagram showing an IZ curve.

FIG. 12 (1) shows an I-Z curve of a wavelength of green (G) color as a characteristic in a narrow wavelength band. In this figure, the peak of the brightness value showing the maximum brightness can be clearly discriminated. On the other hand, when the wavelength band of the illumination light is not limited in the normal optical system, the obtained IZ curve has a gentle peak mainly due to the influence of the chromatic aberration generated in the objective lens 6. I will end up. In (2) of FIG. 12, a situation in which the positions showing the maximum brightness at the respective wavelengths of red (R), green (G), and blue (B) are different due to the chromatic aberration of the lens, and as a result, the combined white color The I-Z curve of light shows a situation where the peak is gentle.

Therefore, a method of obtaining a steep I-Z curve by using a narrow wavelength band by inserting a wavelength filter in the light source or the image pickup means is used. In such a case, the constructed three-dimensional image will be more accurate, but since color information cannot be reproduced, it becomes difficult to identify defects such as flaws and adherents on the sample surface that can be identified by color. It causes problems and cannot be used for visual inspection of samples.

Further, when an image is picked up in a certain selected wavelength band, the reflectance in the used wavelength band is low depending on the sample, the signal level is lowered, and the S / N ratio is deteriorated. It may not be possible to construct a good three-dimensional image.

The present invention has been made in view of the above circumstances, and a steep I-Z curve can be obtained and deterioration of the S / N ratio can be prevented even when normal illumination is used. It is an object of the present invention to provide a scanning confocal microscope capable of obtaining a highly accurate three-dimensional image and reproducing color information on the surface of a sample.

[0014]

In order to solve the above problems, the present invention selects a luminance signal in an optimum wavelength band from a plurality of luminance signals having different wavelength bands of light photoelectrically converted and output by an image pickup means. The scanning confocal microscope is provided with the selecting means and the image constructing means for constructing a three-dimensional image or an image with a deep focal depth by using the luminance signal in the optimum wavelength band.

According to the present invention, in the scanning confocal microscope according to the above-mentioned invention, the image constructing means indicates the maximum luminance of the luminance signal in the optimum wavelength band and the maximum luminance thereof for each pixel of the image pickup device of the image pickup means. Z which is the position in the optical axis direction to be given
It is a scanning confocal microscope that constructs a three-dimensional image or an image with a deep depth of focus by using the directional position and using it.

According to the present invention, in the scanning confocal microscope according to the above-mentioned invention, the selecting means selects the luminance signal in the optimum wavelength band for each pixel, and the image constructing means selects between the respective wavelength bands. It is a scanning confocal microscope that corrects the Z-direction position, which is the position in the optical axis direction, based on chromatic aberration information.

Further, the present invention is the scanning confocal microscope according to the invention described above, wherein the imaging means has a color imaging element.

Further, the present invention is the scanning confocal microscope according to the above-mentioned invention, comprising a plurality of color filters, and switching the color filters to obtain a plurality of luminance signals having different wavelength bands of light. It is a microscope.

Here, the "wavelength band of light" refers to a band of wavelengths forming a main component of the wavelengths forming the light, and "different wavelength bands of light" means the light The wavelength bands that give the maximum brightness do not overlap, and even if there are some overlapping parts in the wavelength bands of the mutual light, the overlapping ratio is small.

[0020]

1 is a diagram showing the configuration of a scanning confocal microscope according to a first embodiment of the present invention.

In this figure, parts having the same functions as those in FIG. 11 are designated by the same reference numerals, and detailed description thereof will be omitted.

Illumination light emitted from the light source 1 passes through a collimator lens 2 and a half mirror 3 and illuminates a rotating disk 4 having a plurality of pinholes formed therein. The rotating disk 4 is attached to the rotating shaft 5a of the motor 5 and rotates at a predetermined rotation speed, and the illumination light that has passed through the plurality of pinholes formed in the rotating disk 4 is sampled by the objective lens 6. Imaged on 7.

The reflected light from the sample 7 again passes through the objective lens 6 and the pinhole of the rotary disk 4 and the half mirror 3, and is condensed by the condenser lens 8 to form a color image pickup section 19.
Is imaged. The computer 20 has a color imaging unit 19
The luminance signals (R, G, B signals) output from the color image pickup device provided in are captured and subjected to image processing, and an image or a calculation result is displayed on the monitor 11.

In response to a command from the computer 20, the Z drive unit 12 relatively moves a part or the whole of the optical system and the sample in the optical axis direction to change the focus position of the sample. In this embodiment, only the objective lens 6 is moved.

Further, the wavelength selecting section 2 is provided in the computer 20.
The signal from No. 1 is input, and the wavelength band used when constructing a three-dimensional image can be selected from the RGB luminance signals read from the color image pickup device of the color image pickup unit 19. In this embodiment, the wavelength selection unit 21
Is configured separately from the computer 20, but is not limited to this, and may be configured as software operating on the computer 20.

FIG. 2 is a diagram illustrating a method for constructing a three-dimensional image of a sample by the scanning confocal microscope of the present invention.

FIG. 2A shows a plan view and a sectional view of the sample 7, which has a three-dimensional (three-dimensional) structure in which the hatched portion in the plan view is a convex portion. Two points in this figure (a,
A) A case of obtaining the height ΔZ between will be described as an example.

FIG. 2B shows IZ curves of color luminance signals R, G and B signals at respective pixels corresponding to two points (a, a) of the color image pickup device. .
An accurate ΔZ can be obtained by selecting any one of the R, G and B signals and obtaining the difference (ΔZ) in the Z direction between the peak positions.

In the case of the example shown in the figure, the R signal is large and the G and B signals are small. This is due to the influence of the color of the surface of the sample 7, and when the surface of the sample 7 is reddish, the G and B signals become small in this way. Therefore, in the case of obtaining ΔZ using G signal for this sample 7, R
The S / N ratio is low as compared with the case of using a signal, and the accuracy of ΔZ deteriorates. This phenomenon occurs not only when the R, G, and B signals are used, but also when the color filter is inserted in the light source and the illumination is narrowed down to a narrow band.

Therefore, in this embodiment, the wavelength selection unit 2
1 selects one of the R, G, and B signals, and the computer 20 is configured to use the selected signal to construct a three-dimensional image.

FIG. 3 is a flow chart showing a schematic operation procedure of the wavelength selecting section 21.

The wavelength selecting section 21 creates an IZ curve for each R, G, B signal for a predetermined pixel of the color image pickup device of the color image pickup section 19 or a pixel in a predetermined area (S).
1). Then, the peak brightness of these I-Z curves is obtained for each R, G, B signal (S2), and the R, G, B signals that give the maximum peak brightness are selected (S3). Then, the selected signal is instructed to the computer 20 (S4).

Here, the signal is selected based on the peak luminance, but the present invention is not limited to this example, and the signal is selected from a predetermined algorithm based on the sharpness index such as peak luminance, S / N ratio, and full width at half maximum. Alternatively, the operator may judge the color of the sample 7, select a signal, and specify and input the signal to the wavelength selection unit 21.

FIG. 4 is a flow chart showing a schematic operation procedure of the computer 20.

The computer 20 uses the signal selected by the wavelength selector 21 to find the Z-direction position that gives the maximum brightness for each pixel (S11), and constructs a three-dimensional image based on this position information (S12). ). Then, based on this information, the distance between the two designated points (a, a) is calculated (S13).

FIG. 2C shows the selected best S / N.
It shows a state in which ΔZ is calculated based on the R signal giving the ratio.

Next, the computer 20 starts to create a color image. First, the maximum brightness, which is the peak brightness of the R, G, and B signals, is obtained for each pixel (S14). Then, at the position in the Z direction that gives the maximum brightness of the selected signal (R signal in this embodiment), the data is configured so that the other signals (G and B signals) also have the maximum brightness (S15). Color information is created using the maximum brightness of the G and B signals and reflected in the three-dimensional image (S16).

As described above, according to the scanning confocal microscope of the present embodiment, an appropriate wavelength band can be selected corresponding to the sample 7, so that a signal with a good S / N ratio can be obtained. It is possible to accurately construct a three-dimensional image using, and to display the three-dimensional image in color. As a result, it is possible to simultaneously perform three-dimensional measurement of the sample 7 and visual inspection such as defect detection.

In this embodiment, the computer 20
The color information was reflected in the three-dimensional image by the process of
As a configuration capable of switching to a non-confocal observation optical path in the optical system, it is possible to reproduce a color close to an actual color by reflecting a non-confocal image.

For example, in parallel with or at a timing different from the operation for constructing a three-dimensional image using the luminance signal in the selected wavelength band, a color image (non-confocal focus) without a rotating disk 4 is used. It is also possible to collect a (image) corresponding to the Z direction position and combine the image information to reflect the color image in the three-dimensional image.

Further, in the present embodiment, the color image pickup device of the color image pickup unit 19 outputs an RGB signal as an example, but the present invention is not limited to this example, and an RGB signal from an output signal of the color image pickup device is used. It is possible to apply the present invention as long as can be extracted. Therefore, YUV output or NTSC output color image pickup devices may be used, and RGB signals may be extracted and used by using these outputs.

FIG. 5 is a view showing the arrangement of a scanning confocal microscope according to the second embodiment of the present invention. In this figure, parts having the same functions as those in FIGS. 1 and 11 are designated by the same reference numerals, and detailed description thereof will be omitted.

The scanning confocal microscope of the present embodiment has a structure in which a monochrome image pickup device is used for the image pickup section 9 and a color filter unit 24 is inserted in the optical path after the light source 1.
The color filter unit 24 is attached to the rotary shaft 23a of the motor 23 and is rotated by a command from the computer 20.

FIG. 6 is a diagram showing the configuration of the color filter unit 24.

The color filter unit 24 is provided with three filters in the wavelength bands of red, green and blue, and is configured so that one of the filters is inserted in the optical path when it is rotated to a predetermined position. There is.

FIG. 7 is a flow chart showing a schematic operation procedure of the computer 20.

The computer 20 uses the color filter unit 2
4 is rotated to switch the color filter (S21). Then, the luminance signal of the sample 7 is read from the image pickup device of the image pickup unit 9 under the selected narrow wavelength band illumination and stored (S
22). This process is repeated for a predetermined color filter to read the luminance signal from the image pickup device of the image pickup unit 9,
When the processing is completed for the luminance signals of all the predetermined color filters (S23), the Z driving unit 12 is operated to move the objective lens 6 by a predetermined distance (S24), and the objective lens 6 performs this processing within a predetermined range. Repeat until the movement is completed (S25).

Since the necessary luminance signal is read into the computer by the above processing, the same processing as the above (S
A three-dimensional image can be constructed by 1 to S16) (S26).

In this embodiment, since the wavelength band of the illumination light is selected by using the color filter, various wavelength bands can be freely selected as compared with the first embodiment.
Therefore, the number of wavelength bands that can be selected is not limited to three, and for example, many narrow wavelength band filters are prepared in order to make the IZ curve steep, and the wavelength band is finer than the wavelength bands of R, G, and B. Can be selected to improve accuracy. When a large number of wavelength bands are selected finely, as many color filters as the number of wavelength bands are arranged in the color filter unit 24.

As described above, according to the scanning confocal microscope according to the second embodiment, the wavelength band corresponding to the sample 7 can be selected similarly to the first embodiment, so that S / It becomes possible to construct a three-dimensional image with high accuracy using a signal with a good N ratio, and it is possible to reflect the color of the sample 7 in the three-dimensional image although the processing time is longer than that in the first embodiment. As a result, three-dimensional measurement of the sample 7 and appearance inspection such as defect detection can be performed at the same time.

Particularly in the present embodiment, since the width and number of wavelength bands can be freely set and selected,
It is possible to perform measurement using a wavelength band more suitable for the sample, and it is possible to further improve the measurement accuracy.

FIG. 8 is a diagram showing a scanning confocal microscope according to the third embodiment. In this figure, parts having the same functions as those in FIGS. 1, 5 and 11 are assigned the same reference numerals and detailed explanations thereof are omitted.

In this embodiment, the color image pickup section 19 is used as the image pickup means, and the computer 30 uses the color image pickup section 19 as a color image pickup section.
Is configured to input a color luminance signal (R, G, B signals) from a color image pickup device provided in
The computer 30 includes a wavelength selection unit 31 and a chromatic aberration storage unit 3.
2 and are incorporated.

The wavelength selecting section 31 compares the R signal, G signal, and B signal for each pixel or for each designated area from the color luminance signal taken in by the computer 30, and finds the signal having the maximum luminance value. By selecting, for each pixel,
The optimum wavelength band is selected for each corresponding part of the sample 7.

In order to correct chromatic aberration, the chromatic aberration storage unit 32 stores a position correction value which is chromatic aberration information between respective wavelength bands, and the wavelength selection unit 31 uses this position correction value to shift the position due to chromatic aberration. It also has a function to correct the.

In the present embodiment, the wavelength selecting section 31 is realized by software operating in the computer 30,
The chromatic aberration storage unit 32 is realized by a memory in the computer 30, but these functions can be configured by hardware.

FIG. 9 is a diagram illustrating a method for constructing a three-dimensional image of a sample according to the third embodiment.

FIG. 9A is a diagram showing a plan view and a sectional view of the sample 7, which has a three-dimensional (three-dimensional) structure in which the hatched portion in the plan view is a convex portion. Two points in this figure (a,
(C) A case of obtaining the height ΔZ between the two will be described as an example.

FIG. 9B shows the IZ curve of the color luminance signals R, G, B signals at the respective pixels corresponding to the two points (A, C) of the color image pickup device. .
An accurate ΔZ can be obtained by selecting any one of the R, G and B signals and obtaining the difference (ΔZ) in the Z direction between the peak positions.

In the example shown in the figure, the R signal is large and the G and B signals are small at the point A. This is due to the influence of the color of the surface of the sample 7, and when the surface of the sample 7 is reddish, the G and B signals become small in this way. On the other hand, at the point C, the G signal is large and the R and B signals are small. When the surface of the sample 7 is greenish, the R and B signals become small in this way.

Therefore, if the wavelength band to be used for this sample 7 is selected as green in advance and the same green wavelength band is applied to the entire screen, this is the optimum selection for point c. However, for the point a, the peak position of the luminance value is obtained using a signal having a poor S / N ratio, resulting in a decrease in accuracy.

Therefore, in the present embodiment, the wavelength selection unit 3
1 is configured to select any one of R, G and B signals for each pixel.

FIG. 10 is a flow chart showing a schematic operation procedure of the wavelength selecting section 31.

The wavelength selecting section 31 obtains the maximum brightness for each R, G, B signal for each pixel of the color image pickup device and selects a signal used for constructing a three-dimensional image (S31). And
The Z-direction position that gives the maximum brightness of the selected signal is obtained (S32).

In this way, the peak of the brightness value can be obtained by using the signal with the best S / N ratio in all the places in the screen. However, as described above, since chromatic aberration occurs in a normal optical system, an error will occur when the Z-direction distance between peaks in different wavelength bands is obtained.

FIG. 9C is a diagram for explaining the influence of chromatic aberration.

In this figure, since the distance is calculated using the R signal and the G signal, the value is ΔZ1, which is an error that is affected by chromatic aberration by Z _RG as compared with the true value ΔZ. ing.

Therefore, the wavelength selection unit 31 extracts the position correction value, which is the chromatic aberration information between the respective wavelength bands, from the chromatic aberration storage unit 32 (S33). This position correction value corrects the Z-direction position of the R signal and the B signal with reference to the Z position of the G signal. Therefore, when the selected signal is the R signal, the position correction value Z _RG is added to the Z direction position, and when the selected signal is the B signal, the position correction value Z _GB is added from the Z direction position. By subtracting, it can be converted into the Z position of the G signal.

The position correction value has a unique value corresponding to the objective lens 6 to be used, but in the present embodiment, this value is not only one type, but the value may be determined and stored for each pixel. Alternatively, the value may be determined and stored for each predetermined area.

In this way, the Z-direction position obtained from each selected signal is corrected by the position correction value (S34), and then the above-mentioned three-dimensional image construction processing is executed using the Z-direction position (S35).

As described above, according to the embodiment of the present invention, since the wavelength band to be used can be selected for each pixel or each designated area, a signal having a good S / N ratio is always adopted. It becomes possible to construct an accurate three-dimensional image and accurately perform three-dimensional measurement.
Further, similarly to the first embodiment, since the color of the sample can be reproduced, the appearance inspection such as the defect detection can be carried out at the same time.

In the present embodiment, the color image pickup device is used as the image pickup means, and the wavelength band to be used has three wavelength bands determined by the R signal, the G signal and the B signal of the luminance signal. However, not limited to this, for example, as in the case of the second embodiment, a plurality of color filters may be arranged in the optical path and a wavelength band determined by the arranged color filters may be used. In this case, an image must be captured for each wavelength, which requires processing time, but it is possible to measure the three-dimensional shape with higher accuracy.

In this embodiment, for example, the rotary disk 4 having spirally arranged pinholes is used, but the rotary disk having slits may be used.
Further, the present invention does not require rotation, but may be configured by using a mask pattern member capable of controlling transmission and blocking of light by changing a predetermined pattern. As such a thing, for example, the rotating disk 4
For example, a case of using a liquid crystal instead of is applicable.

[0074]

As described above, according to the scanning confocal microscope of the present invention, it is possible to obtain a highly accurate three-dimensional image and also to reproduce the color information on the sample surface.

[Brief description of drawings]

FIG. 1 is a diagram showing a configuration of a scanning confocal microscope according to a first embodiment of the present invention.

FIG. 2 is a diagram illustrating a method for constructing a three-dimensional image of a sample by the scanning confocal microscope of the present invention.

FIG. 3 is a flowchart showing a schematic operation procedure of a wavelength selection unit.

FIG. 4 is a flowchart showing a schematic operation procedure of a computer.

FIG. 5 is a diagram showing a configuration of a scanning confocal microscope according to a second embodiment of the present invention.

FIG. 6 is a diagram showing a configuration of a color filter unit.

FIG. 7 is a flowchart showing a schematic operation procedure of a computer.

FIG. 8 is a diagram showing a scanning confocal microscope according to a third embodiment.

FIG. 9 is a diagram illustrating a method of constructing a three-dimensional image of a sample according to the third embodiment.

FIG. 10 is a flowchart showing a schematic operation procedure of a wavelength selection unit.

FIG. 11 is a diagram showing a configuration of a conventional disc scanning confocal microscope.

FIG. 12 is a diagram showing an IZ curve.

[Explanation of symbols]

1 ... Light source 2 ... Rotating disk 4 6 ... Objective lens 7 ... Sample 9 ... Imaging unit 10 ... Computer 11 ... Monitor 12 ... Z drive unit 20 ... Computer 21 ... Wavelength selection section 24 ... Color filter unit 30 ... Computer 31 ... Wavelength selection unit 32 ... Chromatic aberration storage unit

Claims (5)

[Claims] 1. An illuminating light from an illuminating means is imaged on a sample from a mask pattern member which is changed in a predetermined pattern through an objective lens, and reflected light from the sample is again passed through the mask pattern member from the objective lens. A scanning confocal microscope which is incident on an image pickup means, obtains an observation image of the sample while changing the focal point position on the sample in the optical axis direction, and constructs a three-dimensional image of the sample or an image with a deep depth of focus. In the above, the selection means for selecting a luminance signal in the optimum wavelength band from a plurality of luminance signals having different wavelength bands of light, which are photoelectrically converted and output by the image pickup means, and the luminance signal in the optimum wavelength band are used. An image constructing means for constructing a three-dimensional image or an image having a deep depth of focus, and a scanning confocal microscope. 2. The image constructing means, for each pixel of the image pickup device of the image pickup means, has a maximum luminance of a luminance signal in the optimum wavelength band and a Z direction which is a position in the optical axis direction which gives the maximum luminance. The scanning confocal microscope according to claim 1, wherein a position and a position are obtained and a three-dimensional image or an image having a deep depth of focus is constructed by using the position. 3. The selecting means selects a luminance signal in the optimum wavelength band for each pixel, and the image constructing means determines a position in the optical axis direction based on chromatic aberration information between the respective wavelength bands. The scanning confocal microscope according to claim 1 or 2, wherein the position in the Z direction is corrected. 4. The scanning confocal microscope according to claim 1, wherein the image pickup means includes a color image pickup device. 5. A plurality of color filters are provided, and the color filters are switched to obtain a plurality of luminance signals having different wavelength bands of the light, according to any one of claims 1 to 3. The scanning confocal microscope described.