JPH09304598A - X ray filter and x-ray microscope - Google Patents

Abstract

PROBLEM TO BE SOLVED: To sufficiently remove X rays adjacent to a 'water window' wavelength range, especially on the side of long wavelengths, and make it possible to solve problems about the deterioration of contrast and the metamorphosis of a sample by filling an X-ray transmission window with water to seal it. SOLUTION: The X-ray filter has members 13 and 13' equipped with an X-ray transmission window 12 such as a silicon nitride thin film and a spacer held between the members. The space between the spacer and the members is filled with water 11, which is sealed by a container 14, a lid 15, an O ring 16 and a screw 17. The X-ray microscope in this invention, which is equipped with a filter 24 in this invention that is constituted as mentioned above and Walter mirrors (or oblique-incidence mirrors) 23 and 26, irradiates a sample such as a living organism in a sample container 25 with X rays of a prescribed wavelength range, and an X-ray detection system 27 picks up images. The filter 24 can sufficiently remove X rays adjacent to a 'water window' wavelength range, especially on the side of long wavelengths, which deteriorates contrast and oxidizes and metamorphoses the sample by evolving free-radical oxygen.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to an X-ray filter, an X-ray filter and a Walter mirror (or an oblique incidence mirror).
The present invention relates to an X-ray microscope equipped with.

[0002]

2. Description of the Related Art Recent developments in biotechnological technology largely depend on observation means such as optical microscopes and electron microscopes.
However, although an optical microscope can observe a living biological sample in a liquid, its spatial resolution is limited to the wavelength of visible light, and an electron microscope has a high spatial resolution, but it is necessary to arrange a sample container in a vacuum. In addition, since there is no window material that constitutes the sample container that allows electron beams to pass therethrough, it was not possible to observe the living biological sample contained in the sample container (non-vacuum atmosphere).

Therefore, an X-ray microscope, which has the possibility of observing living biological samples with high resolution, has been attracting attention and has been developed. Then, due to the development of fine precision engineering, the performance of the X-ray optical element for the X-ray microscope is improved,
A line microscope testing machine is being built. In an X-ray microscope, a wavelength region called a “water window” (2.33 to 4.3) in which a contrast due to a difference in X-ray absorption between a biological sample and surrounding water contained in a sample container is best obtained.
X-ray of 37 nm) is used.

Here, 2.33 nm is the absorption edge of oxygen, which is the main component by weight of water, and 4.37 nm is the absorption edge of carbon, which is the main component by weight of most biological materials. As shown in FIG. 3, in the “water window” wavelength region (2.33 to 4.37 nm),
Since the absorption of X-rays by water is small and the absorption by biological substances is large, a good contrast of the biological sample with respect to water can be obtained.

As an X-ray source for an X-ray microscope, a laser plasma X-ray source, a Z-pinch plasma X-ray source, or the like has been developed in place of the conventional electron impact type, and has a laboratory size of high-intensity X-ray.
The radiation source is ready for use. Note that the actual X
The size of the plasma itself, which is the radiation source, is about 100 μmφ. Generally, a zone plate is used as an illumination optical system element and an imaging optical system element for an X-ray microscope. Since the zone plate has a spectral function by itself,
When using X-rays in the “water window” region for sample observation, no particular attention needs to be paid to the spectrum of incident X-rays on the illumination optical system element.

However, since the zone plate has a small numerical aperture and a spectral function, the utilization efficiency of incident X-rays is not good. Therefore, as research progresses, it takes time to image a biological sample with an X-ray microscope that uses a zone plate as an optical element, and in the case of a sample that moves during that time, the image is blurred and clear imaging cannot be performed. The existence of dots has become clear.

Therefore, since the numerical aperture is large and the reflection has almost no wavelength dependence, the Wolter mirror (or the grazing incidence mirror), which has a high utilization efficiency of incident X-rays and is capable of imaging in a short time, is drawing attention. There is. FIG. 6 shows the Walter mirrors 63 and 66.
FIG. 6 is a schematic configuration diagram of a conventional X-ray microscope (example) that uses the as an optical element.

This X-ray microscope includes a plasma X-ray source 61,
Titanium foil filter 62, Wolter mirror 6 for condensing (illumination)
3, a sample container 65, an objective (imaging) Walter mirror 66, and an imager 67.

[0009]

The spectrum S1 of the X-ray 1 emitted from the plasma X-ray source 61, transmitted through the titanium foil filter 62 and condensed by the Walter mirror 63, and incident on the sample in the sample container 65. X-ray 2 spectrum S2
Is shown in FIG. X-ray 2 incident on the sample in the sample container 65
Passes through the X-ray transmission window (0.05 μm thick silicon nitride film) of the sample container 65, and therefore the spectrum S2 has a reduced intensity as a whole, and is remarkable in the region of wavelength 3.1 nm (absorption edge of nitrogen) or less. The strength is reduced.

FIG. 3 shows that X-rays having a wavelength of 4.37 nm or more not only contribute to the contrast of the biological sample with respect to water but also reduce the contrast. Then, from the spectrum S2 in FIG.
It can be seen that the X-ray component of 37 nm or more is not sufficiently removed from the X-ray (X-ray incident on the sample in the sample container 65) 2.

That is, in an X-ray microscope using a conventional filter (titanium foil filter or the like) and a Walter mirror, unnecessary visible light from the X-ray source, ultraviolet light, and a region outside the "water window" wavelength region are used. Although the X-rays of (1) can be removed by a conventional filter (titanium foil filter or the like), the X-rays in the vicinity of the “water window” wavelength region, especially in the vicinity of the long wavelength side cannot be sufficiently removed.

In the region of wavelength 2.33 nm or more, X of water
The X-ray absorption is smaller than that of the biological sample, but the X-ray absorption of water gradually increases with the increase of wavelength, and the X-ray absorption can be ignored from the area beyond the “water window” wavelength range (2.33 to 4.37 nm). (See FIG. 3). Then, if the X-ray component having a wavelength of 4.37 nm or more cannot be sufficiently removed, and as a result, the X-ray absorption of water becomes non-negligible, the contrast of the biological sample with respect to water decreases, and There is a problem that free radical oxygen is generated by the X-ray absorption of water, and the free radical oxygen causes oxidation of a biological substance and denatures the sample.

The present invention has been made in view of the above problems, and is capable of sufficiently removing X-rays in the vicinity of the "water window" wavelength region, particularly X-rays in the vicinity of the long wavelength side. And an X-ray microscope provided with the X-ray filter and the Walter mirror.

[0014]

Therefore, firstly, the present invention provides an "X-ray filter comprising a plurality of X-ray transmission windows, wherein water is filled between the plurality of X-ray transmission windows (claim 1). )"I will provide a. Further, the present invention is secondly "X-ray having at least a plurality of members having an X-ray transmission window, spacers sandwiched between the members, and water filled between the spacers and the members. Filter (claim 2) ".

The third aspect of the present invention is "at least a plurality of members having an X-ray transmission window, spacers sandwiched between the members, water filled between the spacers and the members, An X-ray filter having a member for sealing water (claim 3) ". Further, a fourth aspect of the present invention is "at least an X-ray source, a filter for removing unnecessary visible light, ultraviolet light, long-wavelength X-rays and short-wavelength X-rays from the X-ray source, and a filter that transmits the filter. An illumination optical system for irradiating the sample with the X-rays, a sample container for holding the sample, an image-forming optical system for forming an image of the X-rays transmitted through the sample at a predetermined position, and arranged at the predetermined position. An X-ray microscope having an X-ray detection system, wherein a Walter mirror or an oblique incidence mirror is used as an optical element of the optical system, and a plurality of X-ray transmission windows are provided between the X-ray source and the sample. An X-ray microscope (claim 4) in which an X-ray filter filled with water is arranged between the plurality of X-ray transmission windows.

In a fifth aspect of the present invention, "the X-ray filter includes at least a plurality of members having an X-ray transmission window,
An X-ray microscope (claim 5) according to claim 4, further comprising a spacer sandwiched between the members and water filled between the spacer and the member. A sixth aspect of the present invention is that "the X-ray filter includes at least a plurality of members having an X-ray transmission window, a spacer sandwiched between the members, and a space between the spacer and the members. An X-ray microscope (claim 6) according to claim 4, comprising water and a member for sealing the water.

[0017]

BEST MODE FOR CARRYING OUT THE INVENTION The present inventor has found that the X-ray absorption coefficient of water is large in the vicinity X-ray region on the longer wavelength side than the “water window” wavelength region (2.33 nm to 4.37 nm), and the wavelength is 2.33 nm. Paying attention to the fact that the X-ray absorption coefficient is relatively large even in the vicinity X-ray region on the shorter wavelength side, and the present invention (X-ray filter and X-ray filter and Line microscope).

That is, the X-ray filter of the present invention has a plurality of X-ray transmission windows, and water is filled between the plurality of X-ray transmission windows (claim 1). An example of the X-ray filter of the present invention is shown in FIG. The X-ray filter F of FIG. 1 includes two members (silicon substrates) 13 and 13 ′ each having an X-ray transmission window (0.05 μm thick silicon nitride thin film) 12, and a spacer S sandwiched between the members. Water 11 filled between the spacer and the member and a member for sealing the water (a container body 14, a container lid 15, three O-rings 16, and four screws 17).

The X-rays transmitted through the titanium foil filter (an example of a filter for removing unnecessary visible light, ultraviolet light, long-wavelength X-rays and short-wavelength X-rays from the X-ray source) are X-ray filters F. When it is irradiated with X-rays, the X-rays pass through the two X-ray transmission windows 12 and the water layer (thickness of 3 μm) 11 to show the spectrum S3 of FIG. 4 (X of wavelength 4.37 nm or more).
Most line components are removed).

That is, in FIG. 4, the spectra S2, S
The ratios of the X-ray components having a wavelength of 4.37 nm or more in 3 are 13.4% and 3.4%, respectively, and it can be seen that in the spectrum S3, most of the X-ray components having a wavelength of 4.37 nm or more are removed. Further, the X-ray filter of the present invention has a relatively large X-ray absorption coefficient even in the vicinity region on the shorter wavelength side than the wavelength of 2.33 nm, and can be used also as a filter for removing the X-ray component in the vicinity region (Fig. 5).

Therefore, the X-ray filter of the present invention can sufficiently remove X-rays in the vicinity of the window wavelength range of water, particularly X-rays on the long wavelength side. The X-ray filter of the present invention includes at least a plurality of members having an X-ray transmission window, spacers sandwiched between the members, and water filled between the spacers and the members. Preferred (Claim 2).

With this structure, the layer thickness of the water through which the X-rays pass is changed according to the height of the spacer,
The filter characteristics can be adjusted. The X-ray filter of the present invention includes at least a plurality of members each having an X-ray transmission window, spacers sandwiched between the members, water filled between the spacers and the members, and the water sealed. Member to stop,
It is preferable to have (Claim 3).

With such a construction, the filter characteristics can be adjusted, water leakage is prevented, and X
It is possible to prevent the degree of vacuum from being lowered due to leakage of water in the vacuum atmosphere in which the line filter is arranged. Next, the X-ray microscope of the present invention includes at least an X-ray source, a filter for removing unnecessary visible light, ultraviolet light, long-wavelength X-rays, and short-wavelength X-rays from the X-ray source, and the filter. An illumination optical system for irradiating the sample with X-rays transmitted through the sample container, a sample container for holding the sample, an imaging optical system for forming an image of the X-rays transmitted through the sample at a predetermined position, and an imaging optical system arranged at the predetermined position. An X-ray microscope having an X-ray detection system comprising: a Wolter mirror or an oblique incidence mirror as an optical element of the optical system; and a plurality of X-ray transmissions between the X-ray source and the sample. An X-ray filter having a window and filled with water is arranged between the plurality of X-ray transmission windows (claim 4).

That is, the X-ray microscope of the present invention has a plurality of X-rays.
An X-ray filter including a ray transmission window, water filled between the plurality of X-ray transmission windows, and a Walter mirror (or an oblique incidence mirror) are provided. Therefore, the X-ray microscope of the present invention has a large numerical aperture and has almost no wavelength dependence in reflection, so that the utilization efficiency of incident X-rays is high and imaging can be performed in a short time. While taking advantage of the advantages of the above, the aforementioned problems associated with irradiation of biological samples with X-rays in the vicinity of the “water window” wavelength region, particularly X-rays on the long wavelength side (contrast reduction, denaturation of samples by free radical oxygen) Can be solved.

Also in the X-ray microscope of the present invention, the X-ray filter is filled with at least a plurality of members each having an X-ray transmission window, a spacer sandwiched between the members, and a space between the spacer and the members. It is preferable to have the following water (Claim 5). With such a configuration, it is possible to adjust the filter characteristics by changing the layer thickness of the water through which the X-rays pass according to the height of the spacer.

Also in the X-ray microscope of the present invention, the X-ray filter is filled with at least a plurality of members each having an X-ray transmission window, a spacer sandwiched between the members, and a space between the spacer and the members. And a member for sealing the water,
It is preferable to have (Claim 6). With such a configuration, the filter characteristics can be adjusted, water leakage can be prevented, and a decrease in vacuum degree due to water leakage of the vacuum atmosphere in which the X-ray filter is arranged can be prevented. .

Hereinafter, the present invention will be described in more detail by way of examples, but the present invention is not limited to these examples.

[0028]

Embodiment 1 FIG. 1 is a schematic sectional view showing an X-ray filter F of this embodiment. As described above, the X-ray filter F of FIG. 1 has an X-ray transmission window (0.05 μm thick silicon nitride thin film).
Two members (silicon substrate) 13 and 13 'having 12
A spacer S sandwiched between the members, water 11 filled between the spacer and the members, and a member for sealing the water (container body 14, container lid 15, three O-rings 1)
It has 6 and 4 screws 17).

That is, the X-ray filter F of this embodiment is
An X-ray transparent film (0.05 mm) formed on two silicon substrates 13
a silicon nitride thin film 12 having a thickness of 12 μm and a spacer S sandwiched between the substrates, and a water layer 11 is sandwiched between the container body 14 and the container lid 15 so that water does not leak out. It is sealed with an O-ring 16 and four screws 17. When the X-ray filter F is irradiated with X-rays transmitted through a titanium foil filter (an example of a filter for removing unnecessary visible light, ultraviolet light, long-wavelength X-rays and short-wavelength X-rays from the X-ray source),
The X-rays are two X-ray transmission windows 12 and a water layer (thickness 3 μm) 11
The spectrum S3 shown in FIG. 4 is obtained by passing through, and almost all the X-ray components having a wavelength of 4.37 nm or more are removed.

In the X-ray filter F of this embodiment, a graph of the X-ray transmittance when the thickness of the water layer is variously changed by changing the thickness of the spacer S and disposing the spacer S respectively. Is shown in FIG. When the thickness is 1 μm, the breakage of 4.37 nm or more is poor, and when the thickness is 10 μm, the X-ray passage through the window region of water is poor. In the end, it turns out to be appropriate to use a layer of water with a thickness of the order of 3-5 μm.

Further, the X-ray filter of this embodiment has a wavelength
The X-ray absorption coefficient is relatively large even in the vicinity of wavelengths shorter than 2.33 nm, and it can be used as a filter for removing the X-ray component in the vicinity (see FIG. 5). Therefore, the X-ray filter of the present embodiment can sufficiently remove X-rays outside the window wavelength range of water, especially X-rays on the long wavelength side.

FIG. 2 shows an X-ray source 21, a filter 22 for removing unnecessary visible light, ultraviolet light, long-wavelength X-rays and short-wavelength X-rays from the X-ray source, and the filter 22 transmitted therethrough. X
An illumination optical system 23 for irradiating the sample with rays, a sample container 25 for holding the sample, an imaging optical system 26 for forming an image of X-rays transmitted through the sample at a predetermined position, and an imaging optical system 26 arranged at the predetermined position. 3 is a schematic configuration diagram of an X-ray microscope including the X-ray detection system 27.

In this X-ray microscope, Walter mirrors 23 and 26 are used as optical elements of the optical system, and
The X-ray filter 24 of the present embodiment is arranged between the radiation source 21 and the sample. Since this X-ray microscope is provided with the X-ray filter 24 and the Wolter mirrors 23 and 26 of the present embodiment, "the incident X-ray has a large numerical aperture and reflection has almost no wavelength dependence. Is highly efficient,
It is possible to image in a short time. "
While taking advantage of the advantage of No. 26, the above problems associated with irradiation of a biological sample with X-rays in the vicinity of the “water window” wavelength region, particularly X-rays on the long wavelength side (contrast reduction, Degeneration) can be solved.

[0034]

As described above, the X-ray filter of the present invention can sufficiently remove X-rays in the vicinity of the "water window" wavelength region, particularly X-rays on the long wavelength side. In addition, the X-ray microscope of the present invention has a large numerical aperture and has almost no wavelength dependency in reflection, so that the utilization efficiency of incident X-rays is high and imaging in a short time is possible. X in the vicinity of the "water window" wavelength region while taking advantage of
It is possible to solve the above-mentioned problems (contrast reduction, denaturation of sample by free radical oxygen) associated with irradiation of biological sample with X-rays, especially X-rays on the long wavelength side.

[Brief description of drawings]

FIG. 1 is a schematic sectional view showing an X-ray filter F of an embodiment.

FIG. 2 is a schematic configuration diagram of an X-ray microscope including the X-ray filter of the embodiment.

FIG. 3 is a data diagram showing X-ray absorption characteristics of water and protein.

FIG. 4 is a data diagram showing each X-ray spectrum after passing through various filters.

FIG. 5 is a data diagram showing the respective X-ray transmittances at different water layer thicknesses in the X-ray filter of the example.

FIG. 6 is a schematic configuration diagram of a conventional X-ray microscope (one example).

[Explanation of symbols]

1 X-ray incident on sample container 2 X-ray incident on sample (conventional case) 3 X-ray incident on sample (when the filter of the present invention is used) 11 Water layer 12 X-ray transmission window 13 X-ray transmission window Attached silicon substrate (an example of a member having an X-ray transmission window) 14 container body (an example of a member for sealing water) 15 container lid (an example of a member for sealing water) 16 O-ring (sealing of water) Example of a member to be used 17 Screw (an example of a member that seals water) 24 Example of a filter of the present invention 21,61 Laser plasma X-ray source 22,62 Titanium foil filter (an example of a conventional filter) 23,63 Condensing ( Illumination Walter mirror (an example of an optical element of an illumination optical system) 25, 65 Sample container 25, 65 Objective (imaging) Walter mirror (an example of an optical element of an imaging optical system) 26, 66 Imager (X-ray) Example of detection system)

Claims (6)

[Claims] 1. An X-ray filter comprising a plurality of X-ray transmission windows, wherein water is filled between the plurality of X-ray transmission windows. 2. An X-ray filter comprising at least a plurality of members having an X-ray transmission window, spacers sandwiched between the members, and water filled between the spacers and the members. 3. A plurality of members having at least an X-ray transmission window, spacers sandwiched between the members, water filled between the spacers and the members, and a member for sealing the water. An X-ray filter having: 4. An X-ray source, a filter for removing unnecessary visible light, ultraviolet light, long-wavelength X-rays and short-wavelength X-rays from the X-ray source, and an X-ray transmitted through the filter. An illumination optical system for irradiating the sample, a sample container for holding the sample, an image forming optical system for forming an image of X-rays transmitted through the sample at a predetermined position, and an X-ray detection system arranged at the predetermined position And a Wolter mirror or an oblique incidence mirror is used as an optical element of the optical system, and a plurality of X-ray transmission windows are provided between the X-ray source and the sample. An X-ray microscope having an X-ray filter, which is filled with water, disposed between the X-ray transmission windows. 5. The X-ray filter is at least X.
The X-ray microscope according to claim 4, further comprising: a plurality of members having a line-transmissive window; spacers sandwiched between the members; and water filled between the spacers and the members. 6. The X-ray filter is at least X.
A plurality of members having a line-transmissive window, spacers sandwiched between the members, water filled between the spacers and the members, and a member for sealing the water. The X-ray microscope according to claim 4.