U.S. patent application number 09/758673 was filed with the patent office on 2002-01-17 for arrangement for spectrally sensitive reflected-light and transmitted-light microscopy.
Invention is credited to Borlinghaus, Rolf.
Application Number | 20020005982 09/758673 |
Document ID | / |
Family ID | 7944047 |
Filed Date | 2002-01-17 |
United States Patent
Application |
20020005982 |
Kind Code |
A1 |
Borlinghaus, Rolf |
January 17, 2002 |
Arrangement for spectrally sensitive reflected-light and
transmitted-light microscopy
Abstract
The arrangement for spectrally sensitive reflected-light and
transmitted-light microscopy comprises an illuminating device which
illuminates a sample under study in two dimensions. A scanning
device is provided which respectively directs the light emanating
from a detection region of the sample under study onto a multiband
detector. In one embodiment, the illuminating device is arranged in
such a way that it transilluminates the sample in two dimensions.
In a further embodiment, the illuminating device illuminates the
sample in two dimensions in reflected light.
Inventors: |
Borlinghaus, Rolf;
(Dielheim, DE) |
Correspondence
Address: |
Simpson, Simpson & Snyder, L.L.P.
5555 Main Street
Williamsville
NY
14221
US
|
Family ID: |
7944047 |
Appl. No.: |
09/758673 |
Filed: |
January 11, 2001 |
Current U.S.
Class: |
359/385 ;
359/368; 359/389 |
Current CPC
Class: |
G02B 21/088
20130101 |
Class at
Publication: |
359/385 ;
359/368; 359/389 |
International
Class: |
G02B 021/06 |
Foreign Application Data
Date |
Code |
Application Number |
Jul 17, 2000 |
DE |
200 12 378.5 |
Claims
What is claimed is:
1. Arrangement for spectrally sensitive reflected-light and
transmitted-light spectroscopy, comprising an illuminating device
(1) for illuminating at least one region of a sample (4) in two
dimensions, a scanning device (6) which defines a detection region
in the sample and a multiband detector (7) which receives the light
emanating from the detection region of the sample in temporal
sequence wherein the light is directed by the scanning device
(6).
2. The arrangement according to claim 1, characterized in that the
illuminating device (1) transilluminates the sample (4) in two
dimensions.
3. The arrangement according to claim 1, characterized in that the
illuminating device (1) illuminates the sample (4) in two
dimensions in reflected light.
4. The arrangement according to claim 1, characterized in that a
pinhole is provided upstream of the multiband detector (7), the
detection region being smaller than the pinhole.
5. The arrangement according to claim 2, characterized in that the
multiband detector (7) records at least two data records of the
sample (4) of different spectral compositions.
6. The arrangement according to claim 5, characterized in that the
multiband detector (7) constitutes a slit/detector arrangement.
7. The arrangement according to claim 5, characterized in that the
multiband detector (7) is formed by at least one dichroic beam
splitter (9, 10) which directs the light onto an assigned detector
(11, 12, 13) in accordance with the filter properties.
8. The arrangement according to claim 1, characterized in that the
scanning device (6) rasterizes the region of the sample (4) in such
a way that the light from the plurality of detection regions, which
are substantially smaller than the region of the sample (4), passes
into the multiband detector (7) sequentially.
Description
CROSS REFERENCE TO RELATED APPLICATIONS
[0001] This invention claims priority of a German filed utility
model application DE 200 12 378.5 filed Jul. 17, 2000 which is
incorporated by reference herein.
FIELD OF THE INVENTION
[0002] The invention relates to an arrangement for spectrally
sensitive reflected-light and transmitted-light microscopy of a
plurality of spectral regions of the reflected light or transmitted
light of a sample, in particular for detecting the light coming
from a scanning device in the detection beam path of a
microscope.
BACKGROUND OF THE INVENTION
[0003] Devices for simultaneous detection of a plurality of
spectral regions have been known from practice for a considerable
time, specifically under the designation of "multiband detector".
These are complicated optical arrangements which permit multiple
focusing with the aid of additional optical systems. Such
arrangements require a very considerable space for spectral
multiband detection, and therefore entail a not inconsiderable
overall size. Moreover, a defocusing effect regularly occurs there,
thus entailing the need for permanent refocusing with the aid of
the additional optical system--referred to the respective spectral
region. These devices are principally used in confocal scanning
microscopy (see in this regard: DE 199 02 625.4 and DE 43 30
447).
[0004] Colour filters which permit detection of a plurality of
wavelengths are likewise used in the prior art, DE 198 35 070, for
analysing the light emanating from the sample. In this case, the
sample is illuminated with the aid of a laser via a scanning device
using reflected light.
[0005] It is not possible using the prior art described above in
conjunction with two-dimensional illumination to take sample data
from a sample under study using reflected light or transmitted
light in arbitrary spectral regions, in particular not
simultaneously in a plurality of spectral regions.
SUMMARY OF THE INVENTION
[0006] It is therefore the object of the invention to create an
arrangement in which the problems known from the prior art are
avoided.
[0007] The above object is achieved by an arrangement for
spectrally sensitive reflected-light and transmitted-light
spectroscopy. The arrangement comprises an illuminating device for
illuminating at least one region of a sample in two dimensions, a
scanning device which defines a detection region in the sample and
a multiband detector which receives the light emanating from the
detection region of the sample in temporal sequence wherein the
light is directed by the scanning device.
[0008] It has been realized according to the invention that the
simultaneous detection of a plurality of spectral regions of the
detecting light (transmitted light emanating from the sample) is
directly possible whenever the detecting light is firstly
spectrally spread out, and the spread-out light is subsequently
split out of the dispersion plane. The splitting of the spread-out
light out of the dispersion plane is performed in the way according
to the invention by means of a special optical arrangement, the
light split into spectral regions, or the spectral regions
themselves being detected, specifically simultaneously. It is
important here that the actual splitting into spectral regions is
preceded by spreading out the detecting light such that the
splitting out from the dispersion plane can take place on the
spread-out beam. In any case, there is no need here for multiple
focusing with the aid of additional optics.
[0009] As already previously stated, two optical arrangements are
provided according to the invention, specifically firstly for
spectrally spreading out the detecting light, and secondly for
splitting and subsequently detecting. It is possible to place
upstream of the arrangement for spectrally spreading out the
detecting light a pinhole onto which the incoming detecting light
is focused, it being possible for the pinhole to be placed directly
downstream of a scanning device.
[0010] From there, the divergent beam runs to the arrangement for
spectrally spreading out the detecting light, this arrangement
comprising focusing optics and dispersion means. The dispersion
means can be designed as a prism with regard to a particularly
simple design. In a way which is further advantageous, a focusing
optical system, which can, in turn, comprise a lens arrangement, is
arranged in each case upstream and downstream of the dispersion
means or the prism.
[0011] The light impinging on the prism is focused by the focusing
optical systems into the slit/detector arrangement, which is still
to be explained later, from where the splitting into spectral
regions takes place.
[0012] Particularly with regard to the small overall size,
reflection means for folding back the spread-out light are arranged
downstream of the arrangement for spreading out the detecting
light, it being possible for the reflection means to be a silvered
surface or a mirror. In any case, the folding back of the
spread-out light, which is performed at least once, permits the
overall size of the device as a whole to be small.
[0013] As already previously mentioned, the detecting light can be
focused into the slit/detector arrangement by means of the focusing
optics. This slit/detector arrangement is therefore arranged in the
beam path of the spread-out light and comprises reflecting surfaces
which form slit diaphragms which decompose the spread-out light
into a plurality of light beams and thus into spectral regions,
doing so, on the one hand, by forming a slit and, on the other
hand, by reflection out of the dispersion plane. In other words,
the slit diaphragms serve the purpose of partially passing the
light arriving there--in accordance with the diaphragm width--and,
on the other hand, of reflecting at the reflecting surfaces
provided there, with the result that even given only one slit
diaphragm and two reflecting surfaces (on both sides one reflecting
surface each for the purpose of forming the slit diaphragm)
decomposition into three light beams and thus into three spectral
regions is possible. This splitting is performed out of the
dispersion plane of the spread-out light. Of course, both the light
beam passed at the slit diaphragm and the reflected partial beam
can impinge once again on a slit diaphragm and be decomposed there
further in accordance with the foregoing explanation. The
decomposition into a plurality of partial beams is therefore
possible through multiple arrangement of slit diaphragms with
appropriate reflecting surfaces.
[0014] The split light beams pass directly to detectors, the number
of the detectors corresponding to the number of the light
beams.
[0015] Moreover, it is essential for the slit diaphragms provided
here that they are placed or arranged approximately at the focus of
the spread-out light. The reflecting surfaces of the slit
diaphragms are designed as silvered surfaces or mirrors, it being
possible for the silvered surfaces to be vapour deposited, for
example, in accordance with the substrate material.
[0016] With regard to a concrete configuration of the slit
diaphragms, it is advantageous when the silvered surface is
assigned to a slit diaphragm jaw which forms the slit diaphragm,
and when the slit diaphragm jaw can be set or adjusted or displaced
in its position which defines the slit diaphragm, the region of the
spread-out light which is to be reflected and, if appropriate, the
reflecting angle. Thus, by setting the slit diaphragm jaw it is
possible to fix not only the spectral region of the light beam
passed and the reflected light beam, but also the direction in
which the reflected light beams run. The arrangement of the
detectors is therefore variable, at least in a certain region.
[0017] In concrete terms, the slit diaphragm jaws could each be
designed as a cubic rod with at least a partially silvered surface.
One of the surfaces then serves, at least partially, as reflecting
surface, being in this case the surface bordering on the actual
slit. A solid glass body comes into consideration for producing the
slit diaphragm jaw, and is already capable of offering total
reflection at its surface depending on the type of glass used.
Moreover, glass can be worked easily and has an extremely small
coefficient of thermal expansion, with the result that there is no
need to adjust the arrangement as a function of temperature.
[0018] In a further advantageous way, the slit diaphragm jaws are
designed as a slide with a rotatably driven spindle and with an
appropriate thread. To this extent, the setting of the slit
diaphragm jaws could be performed via actuators which feed and, if
appropriate, rotate the silvered surface of the slit diaphragm jaw.
The slit width and the width of the reflected light, and thus the
respective spectral region can be set by adjusting the position of
the slit diaphragm jaw. Adjustment to permanently positioned
detectors is possible by adjusting the angular position of the slit
diaphragm jaw, and thus of the reflecting angle. The actuators can
be any desired manual operations. Electric motors, in particular
electric motors with micro-drive, can advantageously serve as
actuators.
[0019] Measures for suppressing stray light can be provided within
the slit/detector arrangement; thus, for example, so-called light
traps or stops such as are adequately known from the prior art for
the purpose of suppressing stray light.
[0020] Any conventional detectors can be used as detectors for the
different spectral regions or colours. Thus, for example, it is
also possible to use commercially available CCD sensors.
[0021] With regard to a compact design of the overall arrangement,
it is advantageous when the device for spectrally spreading out the
detecting light, and the slit/detector arrangement are supported by
a single chassis which can be mounted or fixed directly on the
scanning device. The slit/detector arrangement with the slit
diaphragm jaws provided there could be arranged in a housing which
can be handled as an insert. The insert could, in turn, be
adjustable in position for the purpose of setting the angle of
incidence and the dispersion plane of the spread-out light.
[0022] Finally, it is also conceivable for the housing for the
slit/detector arrangement to be largely thermally insulated in
order, specifically, effectively to avoid thermal influences on the
arrangement there. According to the invention, instead of the
arrangement described, it is also possible to use as multiband
detector an arrangement of dichroic beam splitters which are
preferably fitted in a cascaded fashion.
[0023] There are various possibilities for advantageously
configuring the teaching of the present invention and developing it
further. For this purpose, reference may be made, on the one hand,
to the claims dependent on Patent claim 1 and, on the other hand,
to the following explanation of an exemplary embodiment of the
invention with the aid of the drawing. Generally preferred
refinements and improvements of the teaching are also explained in
conjunction with the explanation of the preferred exemplary
embodiment of the invention.
BRIEF DESCRIPTION OF THE DRAWINGS
[0024] The subject-matter of the invention is illustrated
diagrammatically in the drawing and is described below with the aid
of the figures, in which:
[0025] FIG. 1 shows a diagrammatic illustration of an embodiment of
the invention for transmitted light,
[0026] FIG. 2 shows a diagrammatic illustration of an embodiment of
the invention for reflected light,
[0027] FIG. 3 shows a diagrammatic illustration of an embodiment of
the invention for transmitted light with the use of dichroites,
[0028] FIG. 4 shows a diagrammatic illustration of the principle of
the mode of operation of the slit/detector arrangement, and
[0029] FIG. 5 shows a diagrammatic illustration of a possible
arrangement of the components of the slit/detector arrangement.
DETAILED DESCRIPTION OF THE INVENTION
[0030] FIG. 1 shows an embodiment of the invention for transmitted
light. An illuminating source 1 generates a broadband beam which
propagates essentially along an optical axis 2. A condenser 3
shapes the beam for illuminating a region of a sample 4 with
transmitted light. The transmitted light passes through the
objective 5 onto a scanning device 6. The scanning device 6 leads
the light to a multiband detector 7. Placed upstream of the
multiband detector 7 is a pinhole 7a which thereby directs the
light from a detection region of the sample 4, which is smaller
than the diameter of the pinhole 7a, onto the multiband detector 7.
The design of the multiband detector 7 is explained more precisely
in the following description.
[0031] The exemplary embodiment illustrated in FIG. 2 discloses an
arrangement for reflected-light illumination. In this case, the
elements similar to those in FIG. 1 are denoted by the same
reference numerals. An illuminating source 1 generates a broadband
beam which propagates essentially along an optical axis 2. The beam
is injected into the beam path of a microscope via a beam splitter
8. The reflected light passes to the sample 4 via the objective 5.
The detecting light passes back via the objective 5 to the beam
splitter 8, passes the latter and impinges on a scanning device 6.
The detecting light passes from the scanning device 6 to the
multiband detector 7 via the pinhole 7a.
[0032] A further embodiment of the invention is illustrated in FIG.
3. In this case, the elements similar to those in FIG. 1 are
denoted by the same reference numerals. The embodiment likewise
describes a transmitted-light arrangement. Downstream of the
scanning device 6, the detecting light passes through the pinhole
7a to a multiband detector 7 which, in this embodiment, comprises
at least one dichroite 9 or 10, and therefore distributes the
detecting light to detectors 11 to 13 in a spectrally selective
fashion.
[0033] A detailed view of the multiband detector 7, which is
arranged, for example, in the beam path of the detecting light in
FIG. 1, is illustrated diagrammatically in FIG. 4. The multiband
detector 7 comprises means 33 for blocking out a second spectral
region 34, on the one hand, and for further reflection of at least
a portion 30 or 35 of the spectral region not blocked out here, on
the other hand. The second detector 32 is arranged in the beam path
of the blocked second spectral region 34, and a third detector 36
is arranged in the beam path of the spectral region 35 which is
further reflected.
[0034] The multiband detector 7 also comprises means 38, arranged
in the beam path of the spectral region 35 further reflected, for
blocking out a third spectral region 39, the third detector 36
being arranged in the beam path of the blocked third spectral
region 39. Consequently, a total of three spectral regions 29, 34
and 39 are selected and detected with the exemplary embodiment
illustrated here. In accordance with the discussion in the general
description, it is possible to cascade a plurality of means, which
block out and reflect a plurality of spectral regions, and
detectors 26, with the result that it is also possible straight
away to select and detect even more than three spectral regions at
the same time.
[0035] The means 27 for spectral decomposition of the light beam 14
are designed as a prism in the case of the exemplary embodiment
illustrated in FIG. 4. The means 28, 33 and 38 are respectively
designed as a slit diaphragm, a reflecting coating 41 being
provided in each case for the purpose of reflecting at least a
portion of the non-blocked spectral region on a surface facing the
incident light.
[0036] FIG. 5 shows the principle of the mode of operation and, on
the other hand, the basic arrangement of the respective components
within the slit/detector arrangement or the multiband detector
7.
[0037] In the slit/detector arrangement 7, the spread-out beam 17
is led through the slit diaphragms 20 and the silvered surfaces 21
into a plurality of partial beams or spectral regions 70, 80, 90
and out of the dispersion plane 6 to the detectors 70a, 80a,
90a.
[0038] Both the detectors 70a, 80a, 90a and the actuators 22,
designed as electric motors, are shown only diagrammatically in the
illustration in FIG. 5. The same holds for the slit diaphragm jaws
23, which form the slit diaphragms 20, together with the silvered
surfaces 21, which are provided directly on the slit diaphragm jaws
23.
[0039] FIG. 5 shows clearly that portions of the spectrally
spread-out beam 17 running into the slit/detector arrangement 7 are
deflected upwards and downwards to the detectors 90a, 80a. A
further partial beam or spectral region 70 passes the slit
diaphragm 20 and reaches the detector 70a.
[0040] The actuators 22 permit the slit 20 to be set between the
slit diaphragm jaws 23, it being possible to set individually the
spectral regions 70, 80, 90 whose partial beams finally pass to the
detectors 70a, 80a, 90a.
[0041] The splitting of the incident spread-out beam 17 renders it
possible to place the split diaphragms 20 with sufficient accuracy
in the focus of the spread-out beam 17.
[0042] It may be noted, finally, that only two of the total of four
actuators 22 are illustrated in FIG. 5, for the sake of simplifying
the illustration.
[0043] The arrangement according to the invention renders it
possible to analyse the detecting light coming from the sample 4
spectrally. In this case, the slit opening 20 in the spreading-out
plane is traversed linearly and thereby delivers a spectral
intensity distribution of the light currently striking the
multiband detector 7 from the scanning device 6.
[0044] Using the invention, it is possible with the aid of a simple
illuminating source, which is a conventional halogen lamp, for
example, to generate at various wavelengths images of a sample
under study. In other words, it is possible to record a series of
spectral images of a sample under study which can extend over the
entire visible spectrum. In addition, the multiband detector 7 can
be used to record simultaneously a plurality of images in different
spectral sections of the overall spectrum.
[0045] It is explained below how images in different spectral
regions are recorded sequentially. Use is made for this purpose of
the arrangement with transmitted-light illumination, and the
multiband detector 7 is designed as a slit/detector arrangement
(see FIG. 5). The sample 4 is transilluminated with the aid of the
illuminating device 1. The illumination of the sample 4 is selected
in this case in such a way that it illuminates in two dimensions at
least in the region of the sample which is recorded by the
objective 5. The scanning device 6 arranged downstream of the
objective 5 directs the light emanating from the sample onto the
multiband detector 7. The scanning device 6 in this case rasterizes
the sample or a selected region thereof in a punctiform fashion,
with the result that information on the entire sample region under
study passes to the multiband detector 7 in temporal sequence.
Punctiform does not mean that only points of the sample 4 are
rasterized. Rather, punctiform means that very small detection
regions are scanned one after another. The size of the detection
regions is determined, for example, in accordance with the inlet
port of the multiband detector 7 or, in the case of a pinhole 7a,
in accordance with the diameter thereof.
[0046] As already mentioned above, at least two images in a
different spectral position can be recorded in parallel using the
multiband detector 7. Furthermore, the multiband detector 7 can be
set, for example, so as to record sequentially images which differ
in colour. The multiband detector 7 can be set in such a way that
the entire spectrum is traversed. Spectrally spread-out images of
the sample are obtained as a result, and thus so is information as
to which points of the sample are emitting, passing or reflecting
light of a specific position in the spectrum.
[0047] The invention has been described with reference to a
particular embodiment. It goes without saying, however, that
changes and modifications can be carried out without leaving the
scope of protection of the following claims in the process.
[0048] Parts List
[0049] 1 Illuminating source
[0050] 2 Optical axis
[0051] 3 Condenser
[0052] 4 Sample
[0053] 5 Objective
[0054] 6 Scanning device
[0055] 7 Multiband detector
[0056] 7a Pinhole
[0057] 8 Beam splitter
[0058] 9 Dichroite
[0059] 10 Dichroite
[0060] 11 Detector
[0061] 12 Detector
[0062] 13 Detector
[0063] 14 Light beam
[0064] 17 Spread-out beam
[0065] 20 Slit opening
[0066] 21 Silvered surfaces
[0067] 22 Actuators
[0068] 23 Slit diaphragm jaws
[0069] 26 Detectors
[0070] 27 Means for spectral decomposition
[0071] 28 Blocking means
[0072] 29 Spectral region
[0073] 30 Reflected spectral region
[0074] 31 First detector
[0075] 32 Second detector
[0076] 33 Blocking means
[0077] 34 Second spectral region
[0078] 35 Reflected spectral region
[0079] 36 Third detector
[0080] 38 Blocking means
[0081] 39 Third spectral region
[0082] 41 Reflecting coating
[0083] 70 Spectral region
[0084] 70a Detector
[0085] 80 Spectral region
[0086] 80a Detector
[0087] 90 Spectral region
[0088] 90a Detector
* * * * *