EP1656578A1

EP1656578A1 - Optical device and microscope comprising an optical device for the collinear combination of light beams of varying wavelengths

Info

Publication number: EP1656578A1
Application number: EP04766443A
Authority: EP
Inventors: Volker Seyfried
Original assignee: Leica Microsystems CMS GmbH
Current assignee: Leica Microsystems CMS GmbH
Priority date: 2003-08-14
Filing date: 2004-08-06
Publication date: 2006-05-17
Also published as: US20070070348A1; JP2007502443A; DE10337558A1; WO2005019898A1

Abstract

The invention relates to an optical device containing a dispersive element (1) and a projection lens (5), which define a splitting plane (7), on which a location is assigned to each light wavelength. A microstructured element (9) is located on the splitting plane, said element guiding light beams (13, 19), which emanate from different directions and are focused on the locations corresponding to their wavelengths, through the projection lens (5) to the dispersive element (1), which combines the light beams in a collinear manner.

Description

OPTICAL DEVICE AND MICROSCOPE WITH AN OPTICAL DEVICE FOR COLLINAR COMBINATION OF LIGHT BEAMS OF DIFFERENT WAVELENGTH

The invention relates to an optical device which collinearly combines light beams 10 and a microscope with an optical device. To combine light beams of different wavelengths, dichroic beam splitters are usually used in optics. A point light source for a laser scanning microscope and a method for coupling at least 15 two lasers of different wavelengths into a laser scanning microscope are known from German published patent application DE 196 33 185 A1. The point light source has a modular design and contains a dichroic beam combiner, which combines the light of at least two laser light sources and couples it into an optical fiber leading to the microscope. Arrangements based on dichroic beam splitters often have the disadvantage that combining light beams that have a closely spaced wavelength is not possible at all or only with little efficiency, since dichroic beam combiners with an infinitely steep edge characteristic can only be produced theoretically. 25 A beam combining device for semiconductor lasers is known from European Patent EP 0 473 071 B1, which contains both dichroic mirrors and a polarizing beam splitter prism. With the help of the polarizing beam splitter prism

Hf Light beams which have a linear polarization direction lying perpendicular to one another are combined to form a collinear light beam, which has both polarization directions. This method of producing an illuminating light beam from two individual light beams can only be used to a limited extent for microscopy, since the predetermined polarization characteristic of the resulting one

Illumination light beam often limits the experimental conditions too much.

In scanning microscopy, a sample is illuminated with a light beam in order to observe the reflection or fluorescent light emitted by the sample. The focus of an exposure light beam is moved with the aid of a controllable beam deflection device, generally by tilting two mirrors, in an object plane, the deflection axes usually being perpendicular to one another, so that one mirror deflects in the x direction and the other in the y direction. The mirrors are tilted, for example, with the help of galvanometer control elements. The power of the light coming from the object is measured depending on the position of the scanning beam. The control elements are usually equipped with sensors for determining the current mirror position. In confocal scanning microscopy in particular, an object is scanned in three dimensions with the focus of a light beam.

A confocal scanning microscope generally includes a light source, imaging optics with which the light from the source is focused on a pinhole - the so-called excitation diaphragm - a beam splitter, a beam deflector for beam control, microscope optics, a detection diaphragm and the detectors for detecting the detection - or fluorescent light. The illuminating light is often coupled in via the beam splitter, which can be designed, for example, as a neutral beam splitter or as a dichroic beam splitter. Neutral beam splitters have the disadvantage that, depending on the division ratio, a lot of excitation or a lot of detection light is lost.

Hf The fluorescent or reflection light coming from the object reaches the beam splitter via the beam deflection device, passes it, and is then focused on the detection diaphragm behind which the detectors are located. Detection light that does not originate directly from the focus region takes a different light path and does not pass through the detection aperture, so that point information is obtained which leads to a three-dimensional image by sequential scanning of the object. A three-dimensional image is usually achieved by recording image data in layers, the path of the scanning light beam on or in the object ideally describing a meander. (Scanning a line in the x direction with a constant y position, then stopping the x scan and panning to the next line to be scanned by y adjustment and then, with a constant y position, scanning this line in the negative x direction, etc.) , In order to enable image data to be taken layer by layer, the sample table or the objective is moved after scanning a layer and thus the next layer to be scanned is brought into the focal plane of the objective.

In many applications, samples are prepared with several markers, for example several different fluorescent dyes. These dyes can be excited sequentially, for example with illuminating light beams that have different excitation wavelengths. Simultaneous excitation with an illuminating light beam that contains light of several excitation wavelengths is also common. From the European patent application EP 0 495 930: "Confocal microscope system for multicolor fluorescence", for example, an arrangement with a single laser emitting several laser lines is known. At present, such lasers are mostly designed as mixed gas lasers, in particular as ArKr lasers.

A device for adjustable coupling and / or detection of one or more wavelengths in a microscope is known from German published patent application DE 198 42 288 A1.

It is the object of the present invention to provide an optical device which makes it possible, regardless of the direction of polarization and

Hf regardless of the spectral proximity of the wavelengths to collinearly combine light rays.

This object is achieved by an optical device in which a dispersive element and an imaging optical system define a splitting level, in which a location is assigned to each light wavelength and in which a microstructured element is arranged that comes from different directions and corresponds to their wavelength Locates focused light rays via the imaging optics to the dispersive element that collinearly combines the light rays. The invention has the advantage that light beams which contain a continuous spectrum can also be combined; even if wavelengths of one light beam are within the spectrum of the other light beam.

In the event that one of the light beams contains light of several wavelengths, it is provided that this light beam is spatially split before it hits the microstructured element. This can be done with a further dispersive element, for example with a prism or a grating, or with the dispersive element that combines the light emanating from the microstructured element.

The dispersive element can be designed, for example, as a grating or as a prism. The imaging optics could, for example, be designed as lens optics or as mirror optics. In a special variant, the dispersive element and the imaging optics are combined, for example, as a concave mirror grating. The imaging optics can include both cylindrical and spherical optics. The distance between the dispersive element and the imaging optics on the one hand and the distance between the imaging optics and the microstructured element on the other hand preferably corresponds to the focal length f of the imaging optics. If the imaging optics, for example in the form of a lens, have two different main planes or if, for some reason, a lens combination is preferred, the distances are preferably selected accordingly, so that the imaging of the

Hf different wavelengths are carried out telecentrically on the fission level. The imaging optics is preferably a telecentric imaging system, since there is then no parallel offset of the returning light.

In a special variant, the microstructured element has reflecting and transmitting regions. In this variant, the light of a first light beam is focused on the reflecting areas, while the light of a second light beam is focused on the transmitting areas. The microstructured element could, for example, include a photolithographically partially mirrored glass substrate, to which the reflecting and the transmitting regions are applied in strips. The stripe pattern is preferably perpendicular to the direction of splitting of the dispersive element.

In another embodiment, the microstructured element has mirror surfaces of different inclinations. Preferably a lamellar structure of linear, e.g. rectangular planar areas, each of which is mirrored and inclined in different spatial directions, the line direction being perpendicular to the spectral splitting in the splitting plane. The respective flat surface sections are preferably rotated out of the splitting plane about an axis of rotation lying in the splitting plane, the pivot axis advantageously running perpendicular to the direction of the spectral splitting. In another variant, the planar patches are rotated out of the splitting plane about axes of rotation running parallel to the splitting direction. The microstructured element could consist of a correspondingly processed and mirrored glass material. In a preferred embodiment, the microstructured element contains a micro-electromechanical system (MEMS) or a micro-opto-electromechanical system (MOEMS). A microstructured element designed in this way has the additional advantage that the local reflection angles can be changed by applying voltages. A usable MDM mirror array is manufactured, for example, by Texas Instruments.

Hf In another preferred embodiment, the microstructured element contains a microprism array composed of different prisms or an array with zones that have a different refractive index, which would be possible, for example, by suitably polarized lithium niobate in an electrical field. This variant also enables specific control via the electrical field.

The beam combining technique according to the invention can be combined with other beam combining techniques, i. H. For example, additional beams can be combined to beams that have already been combined in advance.

All parts to be moved during the adjustment are preferably motorized, in particular it can be advantageous if the spectrally selective element is movable along the direction of the spectral splitting. Before or after the optical device according to the invention, the light output varying elements can be arranged, for. B. preferably an AOTF. The optical device is preferably manufactured as a mechanical unit, which further components such. B. may include an AOTF or temperature stabilization. With the technique described, it is possible to thread not only a second, but also a third or further light beam onto a first light beam. This is particularly advantageous in connection with the described MEMS / MOEMS actuators.

In a very particularly preferred embodiment, the optical device is used to generate an illuminating light beam in a scanning microscope, in particular in a confocal scanning microscope.

The subject matter of the invention is shown schematically in the drawing and is described below with reference to the figures, components with the same effect being provided with the same reference symbols. Show:

Hf 1 shows an optical device according to the invention,

2 shows a microstructured element,

3 shows a further microstructured element,

4 shows a further microstructured element,

Fig. 5 shows another optical device according to the invention and

Fig. 6 shows another optical device according to the invention.

1 shows an optical device according to the invention with a dispersive element 1, which is designed as a prism 3, and with imaging optics 5, which together define a splitting plane 7, in which a microstructured element 9 is arranged. The microstructured element 9 is designed as a strip-shaped reflecting glass substrate 11, the strips of the strip pattern being aligned perpendicular to the splitting direction of the prism 3. A first light beam 13, which contains light of two wavelengths, is spatially split by the prism 3 and the resulting partial beams 15, 17 are focused by the lens 5 onto a mirrored strip of the glass substrate 11. A second light beam 19 is focused by optics 21 onto a transmitting strip of the glass substrate 11. The locations at which the partial beams 15, 17 and the second light beam 19 strike the glass substrate 11 correspond to their wavelength according to the splitting characteristic of the prism 3. The partial beams 15, 17 reflected by the glass substrate 11 are guided together with the transmitted second light beam 19 via the lens 5 to the prism 3, which collinearly combines the partial beams 15, 17 and the second light beam 19 to form an output light beam 23. The microstructured element 9 has a slight inclination with respect to the optical axis in order to spatially separate the first light beam 13 and the output light beam 23 from one another. Due to the inclination of the microstructured element 9, the output light beam 23 extends at an acute angle out of the plane of the drawing, which is not the case in the figure shown

Hf is recognizable. However, the inclination affects the functioning of the optical device only very slightly.

FIG. 2 shows the microstructured element 9, which has already been mentioned with reference to FIG. 1. The microstructured element 9 is designed as a glass substrate coated in the form of a strip and has regions 25 and transmitting regions 27. The stripe pattern is, as indicated by the double arrow 29, arranged perpendicular to the direction of the spectral splitting of the dispersive element.

3 shows a microstructured element 9 with plane mirror elements 31-43, which have different inclinations. The plane mirror elements 31-43 can be rotated about axes of rotation which lie perpendicular to the spectral splitting direction in the splitting plane. The microstructured element 9 is designed as a micro-opto-electro-mechanical system (MOEMS), so that the respective inclination angles can be changed by applying voltages.

4 shows a microstructured element with microprisms 45-57. The prisms are inclined about an axis of rotation that runs parallel to the spectral splitting direction.

FIG. 5 shows a further optical device according to the invention, which contains a fully reflecting microstructured element 9 which has a lamella structure 59. As already described with reference to FIG. 1, the first light beam 13 strikes the microstructured element 9. The second light beam 19 is focused by the lens 21 onto a first section of the microstructured element. The partial beams 15, 17 meet other partial parts 63, 65, the partial parts 63, 65 having a different inclination than the partial part 61. The inclinations of the partial parts 61-65 are selected such that the partial beams 15, 17 and the second light beam 19 are directed together via the lens 5 to the prism 3, which collinearly combines the partial beams 15, 17 and the second light beam 19 to form an output light beam 23.

Hf FIG. 6 shows a development of the optical device shown in FIG. 5. In this embodiment variant, the second light beam 19 contains light of several wavelengths and is spatially spectrally split by an element 67, which is designed as a further prism 69, into the partial beams 71 and 73, which are focused by the lens 21 onto different locations of the microstructured element 9 , The microstructured element 9 reflects the partial beams 15, 17 and the partial beams 71, 73 together via the lens 5 to form the prism 3, which combines the partial beams 15, 17, 71, 73 into a collinearly combined output light beam 23. The invention has been described in relation to a particular embodiment. However, it is understood that changes and modifications can be made without departing from the scope of the following claims.

Hf LIST OF REFERENCE NUMBERS

1 dispersive element

3 prism

5 imaging optics

7 Split level

9 micro-structured element

11 glass substrate

13 first light beam

15 partial beam

17 partial beam

19 second light beam

21 optics

23 output light beam

25 mirrored areas

27 transmitting areas

29 Direction of spectral splitting

31-43 mirror elements

45-57 microprisms

59 slat structure

61 section

63 section

65 section

67 further dispersive element

69 other prism

71 partial beam

73 partial jet)

Hf

Claims

claims

1. Optical device in which a dispersive element and an imaging optical system define a splitting level, in which a location is assigned to each light wavelength and in which a microstructured element is arranged which transmits light beams coming from different directions and focused on the locations corresponding to their wavelength guides the imaging optics to the dispersive element that collinearly combines the light rays.

2. Optical device according to claim 1, characterized in that the light beams have different wavelengths.

3. Optical device according to one of claims 1 or 2, characterized in that at least one light beam has a plurality of wavelengths.

4. Optical device according to one of claims 1 to 3, characterized in that the dispersive element spatially spectrally splits at least one light beam before it hits the microstructured element.

5. Optical device according to one of claims 1 to 4, characterized in that a further dispersive element spatially splits at least one light beam before striking the microstructured element.

6. Optical device according to one of claims 1 to 5, characterized in that the dispersive element contains a prism.

7. Optical device according to one of claims 1 to 5, characterized in that the dispersive element includes a grating.

8. Optical device according to one of claims 1 to 7, characterized in that the dispersive element comprises the imaging optics.

Hf

9. Optical device according to one of claims 1 to 8, characterized in that the microstructured element has reflecting and transmitting regions.

10. Optical device according to one of claims 1 to 9, characterized in that the microstructured element has mirror surfaces of different inclinations.

11. Optical device according to one of claims 1 to 10, characterized in that the microstructured element contains MEMS (micro-electro-mechanical system) or MOEMS (micro-opto-electro-mechanical system).

12. Optical device according to one of claims 1 to 1 1, characterized in that the microstructured element includes a micromirror array.

13. Optical device according to one of claims 1 to 12, characterized in that the microstructured element contains a microprism array.

14. Optical device according to one of claims 1 to 12, characterized in that the microstructured element has areas with different refractive index.

15. Microscope with an optical device according to one of the

Claims 1 to 14.

16. Scanning microscope, in particular confocal scanning microscope, with an optical device according to one of claims 1 to 14 for generating an illuminating light beam.

Hf