EP0270968B1 - Roentgen microscope - Google Patents

Description

The present invention relates to an X-ray microscope in which the object is illuminated coherently or partially coherently with quasi monochromatic X-ray radiation via a condenser and is magnified in the image plane by means of a high-resolution X-ray objective designed as a zone plate.

( p = Linienzahl, m = Nummer der noch zu erfassenden Beugungsordnung).Such X-ray microscopes are described, for example, in Part IV of the book "X-Ray Microscopy" by Schmahl and Rudolph, Springer-Verlag 1984. On pages 192/202 of this book there is a description of an X-ray microscope in which each imaging element, that is to say, the condenser and the X-ray objective, is designed as a zone plate. Such a zone plate consists of a large number of very thin rings, for example made of gold, which are applied to a thin carrier film (for example made of polyimide). These rings form a circular grid with a radially increasing line density. The zone plates diffract the incident monochromatic X-ray radiation of the wavelength and thus cause an image. Quasi monochromatic radiation here means radiation of a certain bandwidth Δλ, this bandwidth being given in connection with zone plates by the relationship $\text{λ / Δλ ≈ p. m}$

(p = number of lines, m = number of the diffraction order to be recorded).

In such known X-ray microscopes, the contrast in the image is mediated by photoelectric absorption in the object, i.e. structures are imaged which effect an amplitude modulation of the X-rays passing through.

The wavelength range of the X-rays, which is between 2.4 nm and 4.5 nm, ie between the oxygen K edge and the carbon K edge, is particularly suitable.

This area is also known as the water window, since water has a transmission that is about ten times higher than that of organic materials. In this wavelength range, organic materials and thus cells and cell organelles can be examined in a living state.

The resolution achieved so far in X-ray microscopy is about a factor of 10 better than in light microscopy, with a further increase in X-ray microscope resolution by about an order of magnitude being still possible. The limit resolution in X-ray microscopy of amplitude structures will be given by the radiation exposure of the objects to be examined.

It is now the object of the present invention to provide an X-ray microscope which enables examinations, in particular of biological structures, to be carried out with a radiation dose which leads to a lower radiation exposure of the objects than the previously usual methods, without having to accept a deterioration in the image contrast must become.

This object is achieved, starting from an X-ray microscope according to the invention, in that an element is arranged in the Fourier plane of the X-ray objective, which extends over the area affected by the zero or a preselected other order of the radiation refracted by the object and gives the transmitted radiation a phase shift and that the element additionally has an absorbing effect to compensate for the intensities of the different orders, the areas with absorbing effect and with phase shifting effect being distributed over different areas in the Fourier plane of the X-ray objective.

In the X-ray microscope according to the invention, phase-shifting properties of object structures are used to form contrast. The phase-shifting element arranged in the beam path gives the preselected order of the X-ray radiation coming from the object a phase shift with respect to the other radiation coming from the object that does not pass through the element. The phase-shifted and the unaffected radiation components interfere in the image plane and thereby generate a contrast-correct, enlarged picture of the object.

It has proven to be particularly advantageous to give the zero-order X-ray radiation of the radiation coming from the object a phase shift of 90 ° with respect to the orders refracted by the object structures. This can be done particularly easily since the zero-order radiation illuminates a central circular disk in the Fourier plane of the X-ray objective. A suitable design of the phase-shifting element is described in claims 3 and 4.

ausdrücken läßt. Die Größe β beschreibt dabei die Absorption, die mit kürzer werdender Wellenlänge λ der Röntgen-Strahlung kleiner wird. Die Größe δ ist maßgebend für die Phasenverschiebung, die der durchgehenden Röntgenstrahlung erteilt wird. Die Größe δ variiert im allgemeinen nur sehr langsam mit der Wellenlänge. Aus diesem Grunde kann also bei Ausnutzung der Phasenverschiebung durch das Objekt eine deutliche Verbesserung des Kontrastes im Bild erreicht werden.The invention is based on the knowledge that the refractive index n of an element in the X-ray range is composed of two differently acting variables, which is shown schematically by the relationship $\text{n = 1 - δ - iβ}$

expresses. The quantity β describes the absorption, which becomes smaller as the wavelength λ of the X-ray radiation becomes shorter. The size δ is decisive for the phase shift which is given to the continuous X-ray radiation. The size δ generally varies very slowly with the wavelength. For this reason, when the phase shift is used by the object, a significant improvement in the contrast in the image can be achieved.

In particular, images can also be generated with a lower radiation exposure to the object, the contrast of which is no worse than when the amplitude contrast is used with higher radiation exposure.

This consideration also gives the further essential advantage of the X-ray microscope according to the invention. Since the size δ changes only slightly with the wavelength λ, when using the phase shift, the wavelength range of the X-rays can be shifted to shorter wavelengths at which X-ray microscopy has so far not been possible due to the low absorption, ie small β, because of the low contrast that can be achieved in the image was sensibly possible.

Under certain circumstances, it may also be possible not to influence the zero-order X-ray radiation in the phase, but rather higher orders of the radiation refracted by the object. These orders form rings in the Fourier plane of the X-ray objective, so that the phase-shifting element is designed according to claim 5.

zeigt ist mit einer Phasenverschiebung stets auch eine absorbierende Wirkung verbunden. Dies gilt natürlich auch für das bei dem Röntgenmikroskop nach der Erfindung verwendete phasenschiebende Element. Deshalb werden die Intensitäten der in der Bildebene interferierenden Ordnungen der vom Objekt kommenden Strahlung einander angeglichen. Dazu wird die phasenschiebende und die absorbierende Wirkung des phasenschiebenden Elementes auf verschiedene korrespondierende Flächen in der Fourierebene des Röntgenobjektivs verteilt. Die durch diese korrespondierenden Flächen tretende Strahlung wird dabei unabhängig voneinander in Phase und Amplitude beeinflußt und zwar so, daß die Intensitäten der in der Bildebene interferierenden Ordnungen der Strahlung aneinander angeglichen sind.Like the formula for the refractive index n in the X-ray range, namely $\text{n = 1 - δ - iβ}$

shows is always associated with a phase shift also an absorbing effect. Of course, this also applies to the phase-shifting element used in the X-ray microscope according to the invention. Therefore, the intensities of the interfering orders in the image plane of the radiation coming from the object are adjusted to each other. For this purpose, the phase-shifting and the absorbing effect of the phase-shifting element is distributed over different corresponding surfaces in the Fourier plane of the X-ray objective. The radiation passing through these corresponding surfaces is influenced independently of one another in phase and amplitude, specifically in such a way that the intensities of the radiation interfering in the image plane are matched to one another.

Fig. 1

ein Ausführungsbeispiel für den prinzipiellen Aufbau eines Röntgen-Mikroskops nach der Erfindung;

Fig. 2

die Draufsicht auf eine als abbildendes Element verwendete Zonenplatte;

Fig. 3

das im Mikroskop der Fig. 1 enthaltene phasenschiebende Element in Draufsicht;

Fig. 4

eine Draufsicht eines anderen Ausführungsbeispiels für ein phasenschiebendes Element.

The invention is explained below with reference to Figures 1-4 of the accompanying drawings. In detail show:

Fig. 1

an embodiment of the basic structure of an X-ray microscope according to the invention;

Fig. 2

the top view of a zone plate used as an imaging element;

Fig. 3

the phase-shifting element contained in the microscope of FIG. 1 in plan view;

Fig. 4

a plan view of another embodiment for a phase-shifting element.

In Fig. 1, the radiation coming from an X-ray source is designated by (1). For example, a synchrotron or another source described in Part 1 of the book "X-Ray Microscopy" by Schmahl and Rudolph, Springer-Verlag 1984 can be used as the X-ray source.

The x-ray radiation passes through an x-ray condenser (2) and is guided by this to the object (3) to be observed, which is arranged on a central diaphragm (4). The X-ray radiation deflected by the object (3) passes through a high-resolution X-ray lens (5) and is imaged by the latter into the image plane (6).

With (7) the Fourier plane of the lens (5) is designated, in which the decomposition of the radiation passing through the object (3) is found in harmonic Fourier components. This distribution is represented again in the image plane (6) by a Fourier inverse transformation as a real image.

Zone plates such as are shown, for example, in FIG. 2 are advantageously used as imaging elements (2) and (5). This zone plate consists of a large number of rings which are placed on a very thin carrier foil, e.g. are applied from polyimide. The rings are usually made of gold or chrome and have a low layer thickness of approx. 0.1 µm. The rings form a circular grid with a radially increasing line density.

A phase-shifting and / or absorbing element (8) is arranged in the Fourier plane (7) of the objective (5). This consists, as shown in FIG. 3, of a thin carrier film (9) which is contained in a ring (10) and on which a thin layer of phase-shifting material, for example chrome in the form a central circular disc (11) is applied.

As can be seen from FIG. 1, the zero-order X-ray radiation (1) coming from the object (3) penetrates the central circular disk (11). This radiation is given a phase shift of 90 ° with respect to the orders diffracted by the object structures. In the image plane (6) there is interference between the phase-shifted radiation and the uninfluenced radiation and thus a high-contrast, enlarged image of the object (3) is created, which can be captured directly on a photosensitive layer, for example.

If, for example, X-rays of a wavelength λ = 4.5 nm are used and the central circular disk (11) of the element (8) consists of a 0.09 μm thick chrome layer, then a protein structure of 10 nm thickness in water results in the X-ray microscope of FIG. 1 an approximately 20 times better contrast than the usual image in the amplitude contrast.

4 shows an exemplary embodiment of an element (8) used for phase shift and / or absorption, in which a ring (12) made of the appropriate material, for example chromium, is attached to the carrier film (9). This ring gives higher orders of the radiation deflected by the object a phase shift. Which order is to be influenced is determined by the diameter and the width of the ring (12).

Claims (5)

1. An x-ray microscope in which the object is illuminated coherently or partially coherently via a condenser with quasi-monochromatic x-radiation and is imaged enlarged by means of a high-resolution x-ray objective in the image plane, characterized by the fact that in the Fourier plane (7) of the x-ray objective (5) there is arranged an element (8) which extends over the region of the surface acted on by the zero order or some other pre-selected order of the radiation diffracted by the object (3) and imparts a phase shift to the traversing radiation and that the element (8) has an additional absorbing action for equalizing the intensities of different orders, whereby regions having absorbing action and regions having phase-shifting action are distributed on different surfaces in the Fourier plane (7) of the x-ray objective (5).
2. An x-ray microscope according to Claim 1, characterized by the fact that the element (8) consists of a layer (11, 12) applied on a support foil (9).
3. X-ray microscope according to Claim 2, characterized by the fact that the layer (11) applied to the support foil (9) has the form of a central circular disk (11) having a thickness such that the x-radiation of zero order passing through it experiences a phase shift of 90°.
4. X-ray microscope according to Claim 3, characterized by the fact that the layer (11) applied to the support foil (9) consists, in the case of an x-ray wave length λ = 4,5nm, of a chromium layer of a thickness of 0,09µm.
5. X-ray microscope according to Claim 2, characterized by the fact that the layer (12) applied to the support foil (9) has an annular form which imparts a phase shift to the radiation of the n th order (|n| ≧ 1) diffracted by the object (3).

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