JPH07167999A - Image focusing type x-ray microscope - Google Patents

Abstract

PURPOSE:To provide an image focusing type X-ray microscope capable of enlarging an observation area. CONSTITUTION:In order to enlarge an observation area, an image focusing type X-ray microscope has jogging mechanism 10 for displacing a sample holder 3 accommodating a sample capsule 8 charged with an observation sample, a driving circuit part 11 for driving the jogging mechanism 10, an X-ray driving circuit part 14 for driving an X-ray generator 1, a control circuit part 12 for taking in X-ray image data obtained by an image pickup device 5 and controlling X-ray outgoing timing, image pickup timing and displacement timing, and a memory data processing part 13 for storing plural X-ray image data taken in by the control circuit part 12 and executing image processing to these X-ray image data so as to synthesize images, and the like.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to an X-ray microscope, and more particularly to an imaging X-ray microscope having an image processing function.

[0002]

2. Description of the Related Art In response to the increasing demand for high-magnification observation of living bodies in medicine, biotechnology, etc., a wavelength of 2 to 5 nm
An X-ray microscope using a soft X-ray of a certain degree has been noticed and developed. A conventional X-ray microscope using such soft X-rays has a structure whose outline is shown in FIG.

In this conventional X-ray microscope, the condenser optical system 2 for X-ray focusing, the sample holder 3, the imaging optical system 4, and the image pickup device 5 are arranged so that the output optical axis of the X-ray generator 1 is the same. It has the structure arrange | positioned in series on the top. Here, the optical path length of the X-ray optical system from the X-ray generator 1 to the imaging device 5 is about 2
m. The entire optical system is housed in a vacuum chamber 7 having an exhaust system 6.

In the conventional example, after the sample capsule 8 loaded with the observation sample is set in the sample container 3, the exhaust system 6 is operated to evacuate the vacuum chamber 7 to a vacuum degree of 4. Observation is performed while maintaining the pressure at 8 × 10 to ² Pa or less. The soft X-ray beam emitted from the X-ray generator 1 is converged by the condenser optical system 2 and passes through the sample capsule 8 set in the sample container 3. The passed soft X-rays are
An image is formed on the image pickup device 5 by the image forming optical system 4, and an observation image is given to the monitor device 9 outside the vacuum chamber 7.

In the condenser optical system or the imaging optical system of the X-ray microscope used here, many X-ray optical elements relating to the convergence of soft X-rays are conceivable. Among them, among them,
Attention has been focused on a zone plate utilizing diffraction, a multilayer film reflecting mirror, and an oblique incidence reflecting mirror.

[0006] In particular, as an X-ray optical element of an X-ray microscope for living body observation using soft X-rays having a wavelength of about 2 to 5 nm, the zone plate is considered in view of high X-ray focusing efficiency and high resolution in this wavelength range. It is considered promising to use any one of a Wolter type grazing incidence reflecting mirror, a multilayer film grazing incidence reflecting mirror, or a combination thereof.

[0007]

However, it is difficult for the X-ray optical element as described above to have a larger field of view than that of the visible light optical element. For example, in the Walter type oblique incidence mirror, the magnification is 40 times and the resolution is 0.1 to 0.05 μm.
Then, the visual field diameter is about 10 μm.

Further, in the zone plate, the field diameter cannot be accurately defined by the magnification, resolution and contrast, but since the zone plate is an X-ray optical element utilizing diffraction, it is necessary to make the X-ray monochromatic. Therefore, it is necessary to insert a pinhole of about 5 to 20 μm on the front surface of the sample to achieve monochromaticity. This is the apparent field stop. Therefore, it can be said that the visual field diameter of the zone plate is about 5 to 20 μm of the pinhole diameter.

In the visual field of the above degree, even if the resolution is higher than that of the optical microscope and a minute area of the living body can be seen, the organelle imaged in the minute area remains in the biological sample to be observed. It is difficult to observe where it is located and how it works.

An object of the present invention is to provide an imaging X-ray microscope using an X-ray optical element having a small field of view, which can enlarge the observation region.

[0011]

SUMMARY OF THE INVENTION The above-mentioned object is to use an imaging X-ray microscope in order to match a predetermined observation position of the sample with a position conjugate with the X-ray imaging position. At least one of the X-rays transmitted through the holding table held at a predetermined position, the site at the observation position of the sample, and the fluorescent X-rays generated from the site is detected to perform the observation. X that outputs a signal indicating the X-ray image at the position
X-ray imaging means, an imaging optical system for forming an X-ray image at the observation position on the imaging surface of the X-ray imaging means, moving means for displacing the holding table to change the observation position, and controlling the moving means. Then, while displacing the holding table quantitatively in a predetermined direction and amount,
The X-ray imaging means is controlled so that imaging is performed every time the holding table is displaced, and a signal indicating the obtained X-ray image is accepted,
The image data obtained based on the signal indicating the X-ray image, the control means for outputting together with the observation position information associated with the quantitative displacement, and the plurality of image data and the observation position information output from the control means, And an image processing means for combining and displaying a part or all of them to display the image-forming X-ray microscope.

[0012]

In the imaging X-ray microscope to which the present invention is applied,
The moving unit changes the observation position by displacing the holding table that holds the sample, and the X-ray imaging unit acquires the X-ray image at the observation position.

The control means controls the moving means and the X-ray image pickup means to pick up a plurality of X-ray images detected each time the sample is displaced quantitatively, and fetches a signal showing the X-ray images.
The control means further converts this signal into image data and outputs the image data together with information indicating the observation position at which the image data was acquired.

The image processing means receives a plurality of image data corresponding to the X-ray images obtained at a plurality of observation positions, output from the control means, synthesizes the image data,
indicate. In the case of combining, for example, a plurality of image data are connected according to the positional relationship of the observation positions where they are acquired, combined and displayed in one X-ray image.

According to the above means, it is possible to enlarge the observable region in the X-ray microscope having a small field of view.

[0016]

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS An embodiment of an imaging X-ray microscope to which the present invention is applied will be described with reference to the drawings. Here, members having the same structure and function as those in the conventional example shown in FIG. 4 are designated by the same reference numerals.

As shown in FIG. 1, the X-ray microscope of this embodiment has an X-ray generator 1 for emitting X-rays and a sample holder for emitting the X-rays as a mechanism for detecting an X-ray image. Condenser optical system 2 that converges on the observation position in the observation sample loaded in the sample capsule 8 housed in 3, an imaging optical system 4 that forms an image of X-rays that have passed through the observation sample, and the formed observation sample Imaging device 5 for capturing an X-ray image at the observation position of the, and vacuum container 7 for accommodating these optical system components.
And an exhaust system 6 for exhausting the inside of the vacuum container 7. Here, the optical path length of the X-ray optical system from the X-ray generator 1 to the imaging device 5 is about 2 m.

Condenser optical system 2 and image forming optical system 4
Examples of the X-ray optical element used in the above include, for example, the zone plate, the Walter type grazing incidence reflecting mirror, and the
Any of the multilayer grazing incidence reflecting mirrors or a combination thereof is used.

As the X-ray generator 1, one that generates soft X-rays having a wavelength of about 2 to 5 nm can be used, and in particular, a pulse laser is applied to a target such as tungsten,
A laser plasma X-ray source capable of emitting pulsed X-rays is suitable.

The image pickup device 5 is for converting an X-ray image of the observation region formed by the image forming optical system 4 into an electric signal, and is composed of a CCD for X-ray or a multi-channel plate and a CCD. The configured one is used.

As the sample holder 3 for observing the living body, a holder having good vacuum separation and good X-ray permeability is used. As such a holder, for example, JP-A-3
-197836, Japanese Patent Application Laid-Open No. 3-197837, Japanese Patent Application Laid-Open No. 3-197838, and Japanese Patent Application Laid-Open No. 3-29
The one described in Japanese Patent No. 5440 is used.

In this embodiment, further, as a mechanism that enables observation in a wide area by changing the observation position on the observation sample, a fine movement mechanism 10 that changes the observation position by displacing the sample holder 3, and a fine movement mechanism. A drive circuit unit 11 for driving 10, an X-ray drive circuit unit 14 for driving the X-ray generator 1 to emit X-rays, and a signal indicating an X-ray image at each observation position obtained by the imaging device 5. And a control circuit unit 12 for converting the signal into image data and outputting the data together with a signal indicating the observation position of the X-ray image,
A memory data operation processing unit 13 that stores a plurality of image data output from the control circuit unit 12 together with their observation positions, executes image processing on these image data, and performs combining processing of these images. Have and.

Here, when the position of the sample holder 3 is displaced, the relative positional relationship between the observation sample loaded in the sample capsule 8 housed in the holder 3 and the X-ray optical axis changes, so that the observation is performed. The position where the emitted X-rays pass through the sample changes. Therefore, the observation position on the observation sample changes according to the displacement amount and direction of the fine movement mechanism 10.

Further, in this embodiment, only the fine movement mechanism 10 is mentioned as the moving mechanism of the moving means, but the present invention is not limited to the magnitude of the displacement amount. In addition to the fine movement mechanism 10,
A coarse movement mechanism using mechanical movement may be provided.

The control circuit section 12 further includes a drive circuit section 1
1, by controlling the X-ray drive circuit unit 14 and the imaging device 5, the X-ray emission timing from the X-ray generator 1, the imaging timing of the imaging device 5, and the fine movement mechanism 10 for displacing the sample holder 3. Control the displacement timing of.
The control circuit unit 12, for example, as shown in FIG. 5, achieves various processing means, such as a CPU 30, a ROM 31, and a
It has a RAM 32, an interface 33 that inputs and outputs data to and from other circuit parts, and an address / data bus 34 for transmitting and receiving data between these components.

The present embodiment further includes an input device 20 such as a keyboard for receiving commands relating to various settings for observation of the X-ray microscope of the present embodiment from the outside and outputting them to the control circuit unit 12, and a memory data arithmetic processing unit. And a display device 21 such as a CRT for displaying the image processing result obtained in step 13.

As shown in FIG. 2, the fine movement mechanism 10 includes a sample table 15 provided with an X-ray transmission window 19 for mounting the sample holder 3 and a piezo element 1 for displacing in the X direction shown in the figure.
6, a piezo element 17 that is displaced in the Y direction, and a drive section 18 that receives a drive signal from the drive circuit section 11 and applies a voltage to the piezo element to be driven. Where X
The line-transmissive window 19 has a size of about several mm square and is sufficiently larger than the visual field of the X-ray optical element of this embodiment. The X and Y directions form a plane orthogonal to the X-ray optical axis.

The moving means in the present invention is not limited to the shape of the fine movement mechanism 10 described above. For example, as shown in FIG. 3, the fine movement mechanism 10 may be arranged in the atmosphere via the bellows 22, and the movement means has a movement accuracy similar to the field diameter of the X-ray microscope to which the present invention is applied. If,
The shape, size, arrangement, and driving method are not limited. Further, the displacement direction is not limited to the XY direction, and the displacement may be made in the X-ray optical axis direction to perform observation in the depth direction of the observation sample.

Next, the operation of this embodiment will be described.

In this embodiment, after the sample capsule 8 loaded with the observation sample is set in the sample holder 3, the evacuation system 6 is operated to evacuate the inside of the vacuum chamber 7 to set the degree of vacuum to, for example,
The observation is performed while maintaining the pressure at 4.8 × 10 ² Pa or less.

The control circuit section 12 of this embodiment is initially shown in FIG.
As shown in the flowchart of FIG. 2, the number of X-ray images to be captured, the area area to be observed, the position, and the like of the loaded observation sample are set based on an external command received by the input device 20, and the drive circuit unit is set. 11 is controlled to perform a predetermined initial setting such as the initial setting of the position of the fine movement mechanism 10 (step 100). Here, the required number of X-ray emission, the amount of displacement of the fine movement mechanism 10, the displacement direction, and the number of displacements are calculated and set from the number of input X-ray images, the area of the observation region, and the like.

Next, the control circuit section 12 controls the X-ray drive circuit section 14 to cause the X-ray generator to emit X-rays, and the X-ray emission causes X-rays at the observation position obtained by the image pickup device 5. The data signal of the line image is fetched (step 102). That is, the soft X-ray beam emitted from the X-ray generator 1 is converged by the condenser optical system 2 and passes through the sample capsule 8 set in the sample container 3. The passed soft X-rays are
An image is formed on the image pickup device 5 by the image forming optical system 4. Vacuum tank 7
The control circuit unit 12 installed outside fetches the data signal of the X-ray image obtained by the imaging device 5.

The control circuit section 12 further receives the X
The data signal of the line image is converted into image data, and the converted image data is output to the memory data calculation processing unit 13 together with information indicating the observation position where the data was obtained. The memory data arithmetic processing unit 13 stores the image data and the position information (step 104).

Next, it is judged whether the number of X-ray images set in step 100 has been acquired (step 10).
6). If not acquired, the control circuit unit 12 controls the drive circuit unit 11 to drive the fine movement mechanism 10 in order to displace the observation position on the observation sample with a preset displacement direction and displacement amount. (Step 108) After that, the procedure returns to Step 102, and an X-ray image at a new observation position is acquired.

If the set number of X-ray images have been acquired, the process proceeds to the final step 110. In step 110, the memory data arithmetic processing unit 13 uses the plurality of X-ray image data stored therein to execute image processing such as combining and laminating these X-ray images, and the display device 21.
To display. If necessary, the input device 20 receives a command from the outside, and executes various other image processes such as boundary line extraction and contrast addition.

The operation described above, in particular, the synthesizing process executed by the memory data operation processing unit 13 using a plurality of X-ray image data will be described more specifically with reference to FIG. Here, a laser plasma X-ray source is used as the X-ray generator 1, and the imaging optical system 4 has a resolution of 0.2 μm and a field of view of 10 mm.
The case where a μm Walter element is used will be described.

The control circuit section 12 emits pulse X-rays by the X-ray drive circuit section 14 for driving the laser plasma X-ray source 1, the image pickup timing of the image pickup apparatus 5 (timing of picking up image pickup data), and the fine movement. By interlocking with the displacement timing for driving the mechanism 10, imaging and displacement operations are executed in accordance with each X-ray pulse (step 10 in FIG. 6).
2-108).

That is, the field of view 210 of the imaging optical system 4 described above.
7A, the size of the largest square rectangular field 211 included therein is a square with one side L of about 7 μm. Therefore,
As shown in FIG. 7A, when the object 200 to be observed is larger than these visual fields, an X-ray image through a visual field of maximum 7 μm square can be obtained by imaging with one X-ray pulse. So I can't get the whole picture.

In the present embodiment, the control circuit section 12 is used to obtain an X-ray image of the observation object 200 as shown in FIG.
As shown in (B), the sample stage 15 on which the sample holder 3 loaded with the observation object 200 is placed is raster-scanned by the fine movement mechanism 10 in the X-axis direction and the Y-axis direction according to the arrows to change the observation position. A total of 9 X-ray images of 3 frames x 3 frames are acquired. Here, each circle in the drawing indicates a visual field 210 for each displacement operation, and one frame means a rectangular visual field 211 obtained from the visual field diameter 210. Reference numeral 212 indicates the center of the field of view 210, and the X and Y directions shown in the drawing form a plane that is perpendicular to the X-ray beam.

In the present embodiment, further, imaging is performed at each displacement operation, that is, every time the observation position changes.
The X-ray image obtained through each field of view 210 is converted into an electric signal by the image pickup device 5, and is converted into digital image data,
It is sent to the memory data arithmetic processing unit 13 by the control circuit unit 12.

The memory data arithmetic processing section 13 is configured to
The X-ray images of one sheet are stored as digital images, and further subjected to image processing to bond them together. That is, as shown in FIG. 7C, the acquired X-ray images are displayed adjacent to each other in correspondence with the relative relationship of the regions where they are imaged, so that a 21 μm square (L = 7
The X-ray image of the area (μm) is obtained and displayed as an image (step 110 in FIG. 6).

The present invention is described in this embodiment.
The number of X-ray images captured, the displacement amount of the fine movement mechanism 10, the displacement direction, etc. are not limited. In the present invention, these amounts are determined by the observation area, the size of the field of view of the X-ray optical system used, and the required roughness of observation, that is, the distance between two consecutive observation positions. be able to.

As the process of synthesizing the image data of the X-ray image,
Further, when synthesizing the image data, the matching state of the data at the overlapping boundary line between the two adjacent image data is checked, the error in the displacement operation is obtained, and the sample holder 3 is displaced based on the error. The operation of the fine movement mechanism 10 may be controlled.

This combining processing method will be described with reference to FIG. Here, the two circles shown are two pieces of X-ray image data acquired continuously. Field of view 210, 2
10 'is a physically limited visual field in the present embodiment, and rectangular visual fields 211 and 211' capture an X-ray image,
It is a field of view of a predetermined size provided when image data is obtained. Here, the field of view for forming such image data does not necessarily have to be a square. Any polygonal shape such as a hexagon or an octagon may be used as long as it is smaller than the original physical field diameter 210 and the data contained therein can be used as much as possible.

Numerals 213 and 213 'indicate a rectangular field of view 21.
The control circuit unit 12 controls the operation of the fine movement mechanism 10 so that these boundaries are overlapped with each other when performing the combining process.

In the present embodiment, if there is an error in the operation of the fine movement mechanism 10, the data at the boundary line 213,
It does not match the data at the boundary line 213 '.

Therefore, the memory data arithmetic processing section 13
Using the entire data of the field of view diameter 210 ′, a boundary line that matches the boundary line 213 is searched for, and the position of the found boundary line in the field of view diameter 210 ′ is calculated. The control circuit unit 12 is
The calculation result is accepted and used to drive the drive circuit unit 1.
1 is further controlled to correct the displacement error.

Further, by comparing the signal intensities in the same portion of the observation sample on the boundary line obtained as described above, the signal intensities (gray scale) in these two images can be standardized. Here, the signal intensity means the transmission of X-rays at the observation sample portion at the observation position,
Depends on reflectance. Further, if this scaling is continuously executed, a composite image composed of a plurality of X-ray images using one scaled signal intensity scale can be obtained.

Further, various kinds of image processing can be executed on the X-ray image acquired as described above.
For example, it is possible to perform boundary line extraction, feature extraction, pattern recognition, contrast addition and emphasis, color addition, etc. of the target object.

Further, it is possible to set a large area to be observed, acquire a large number of X-ray images, and use these to synthesize and display one whole image covering the entire observation area. However, in such a case, it is difficult to display all the detailed features and the like in a single image. Therefore, by further providing an input means such as a mouse, the region of interest is specified, and it corresponds to the specified part. Based on the image data
Only that portion may be enlarged and displayed at a predetermined magnification.

According to the present invention, the observable region of the X-ray microscope is enlarged by synthesizing a plurality of X-ray images obtained each time the sample is displaced to form one image. Therefore, it is possible to more clearly know the mechanism of the biological sample larger than the field of view of the X-ray microscope.

[0052]

According to the present invention, by controlling the moving means and the X-ray imaging means, an X-ray image is acquired each time the sample is displaced, and a plurality of the obtained X-ray images are combined. ,
It is possible to enlarge the observation area of the X-ray microscope.

[0053]

[Brief description of drawings]

FIG. 1 is a block diagram showing an embodiment of the configuration of an imaging X-ray microscope to which the present invention is applied.

FIG. 2 is a block diagram showing an example of a configuration of a moving unit in an imaging X-ray microscope to which the present invention has been applied.

FIG. 3 is a block diagram showing another embodiment of the configuration of the imaging X-ray microscope to which the present invention is applied.

FIG. 4 is a block diagram showing a configuration of a conventional imaging X-ray microscope.

5 is a block diagram showing a configuration of a control circuit unit in the embodiment of FIG.

FIG. 6 is a flowchart showing the operation of the embodiment shown in FIG.

FIG. 7 is a view showing an imaging X-ray microscope to which the present invention is applied,
Explanatory drawing explaining the image processing which concerns on the synthesis | combination of the acquired X-ray image.

[Explanation of symbols]

1 ... X-ray generator, 2 ... Condenser optical system, 3 ... Sample holder, 4 ... Imaging optical system, 5 ... Imaging device, 6 ... Exhaust system,
7 ... Vacuum tank, 8 ... Sample capsule, 9 ... Monitor device, 10
... sample stage fine movement mechanism, 11 ... fine movement mechanism drive circuit section, 12 ...
Displacement / imaging timing control circuit section, 13 ... Memory / data arithmetic processing section, 14 ... Laser plasma X-ray source control section, 1
5 ... Sample stand, 16 ... X-direction drive piezo element, 17 ... Y-direction drive piezo element, 18 ... Piezo element drive circuit section, 19
... X-ray transmission window, 20 ... Input device, 21 ... Display device, 22
... Bellows, 200 ... Observation object, 210 ... X-ray optical system visual field, 211 ... Rectangular visual field obtained from visual field 210, 2
12 ... Center of visual field 210, 213 ... One side of boundary line of rectangular visual field.

Claims (8)

[Claims] 1. In an imaging X-ray microscope, a sample is held at a predetermined position in order to match a predetermined observation position of the sample with a position conjugate with the X-ray imaging position. An X-ray image at the observation position by detecting at least one of the X-rays transmitted through the holding base and the site at the observation position of the sample and the fluorescent X-rays generated from the site. X-ray imaging means for outputting a signal indicating that, an imaging optical system for forming an X-ray image at the observation position on the imaging surface of the X-ray imaging means, and movement for changing the observation position by displacing the holding table. The means and the moving means are controlled to quantitatively displace the holding table in a predetermined direction and amount, and the X-ray imaging means is controlled to perform imaging every time the holding table is displaced. The signal representing the X-ray image is received, and the signal obtained based on the signal representing the X-ray image is obtained. A control unit that outputs the image data together with the observation position information associated with the quantitative displacement, and an image that receives a plurality of image data and the observation position information output from the control unit, and combines and displays a part or all of them. An imaging X-ray microscope, comprising: a processing unit. 2. The X-ray image at a plurality of adjacent observation positions according to claim 1, wherein the control unit controls the moving unit and the X-ray imaging unit so that observable visual field regions overlap each other. The image-forming X-ray microscope, which receives the signals indicating the obtained plurality of X-ray images and outputs the image data corresponding to the X-ray images. 3. The image processing means according to claim 2, wherein the storage unit stores a plurality of received image data and an observation position thereof, a processing unit that processes the plurality of image data stored in the storage unit, and a processing unit. The image data output from the control unit, the processing unit displays the boundary lines of the image data having adjacent and overlapping portions, An imaging X-ray microscope, characterized in that 4. The processing unit according to claim 3, wherein the calculated boundary lines overlap each other.
A plurality of image data is combined into one image data, and the display unit, based on the combined image data,
An imaging X-ray microscope characterized by displaying an image. 5. The boundary line indicated by the output signal according to claim 3, wherein the processing unit of the image processing unit outputs a signal indicating a boundary line of each calculated image data. Based on the above, the relative displacement between the two adjacent image data is calculated, the calculated relative displacement is compared with the relative displacement obtained from the observation position information of the two image data, and the displacement error of the moving means is calculated. An imaging X-ray microscope characterized by correction. 6. The X-ray generator according to claim 1, which emits X-rays to irradiate a sample, and the X-rays provided between the sample and the X-ray generator. An imaging X-ray microscope, further comprising a condenser optical system that converges at a position conjugate with the imaging position. 7. The X-ray generator according to claim 6, wherein the X-ray generator generates a pulse X-ray, and the control unit controls the X-ray imaging unit each time the pulse X-ray is emitted. An imaging X-ray microscope, characterized by acquiring an X-ray image. 8. The moving means according to claim 1, wherein the moving means displaces the holding base within a plane orthogonal to the optical axis of the X-ray, and drives the moving mechanism and An imaging X-ray microscope, comprising: a movement control device that controls a displacement amount and a direction.