DE19835070A1 - Arrangement for adjustable wavelength-dependent detection in fluorescent microscope has combination(s) of short and long pass filters producing adjustable band pass filter - Google Patents
Arrangement for adjustable wavelength-dependent detection in fluorescent microscope has combination(s) of short and long pass filters producing adjustable band pass filterInfo
- Publication number
- DE19835070A1 DE19835070A1 DE1998135070 DE19835070A DE19835070A1 DE 19835070 A1 DE19835070 A1 DE 19835070A1 DE 1998135070 DE1998135070 DE 1998135070 DE 19835070 A DE19835070 A DE 19835070A DE 19835070 A1 DE19835070 A1 DE 19835070A1
- Authority
- DE
- Germany
- Prior art keywords
- filter
- short
- arrangement according
- adjustable
- wavelength
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Classifications
-
- G—PHYSICS
- G02—OPTICS
- G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
- G02B21/00—Microscopes
- G02B21/06—Means for illuminating specimens
-
- G—PHYSICS
- G02—OPTICS
- G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
- G02B21/00—Microscopes
- G02B21/0004—Microscopes specially adapted for specific applications
- G02B21/002—Scanning microscopes
Landscapes
- Physics & Mathematics (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- General Physics & Mathematics (AREA)
- Optics & Photonics (AREA)
- Microscoopes, Condenser (AREA)
- Investigating, Analyzing Materials By Fluorescence Or Luminescence (AREA)
Abstract
Description
Die Erfindung betrifft die Hintereinanderschaltung von variablen oder diskreten Kurzpaß- und Langpaßfiltern zur Realisierung eines frei programmierbaren Bandpaßfilters für den Einsatz in der Fluoreszenzmikroskopie, insbesondere in einem Laser- Scanning-Mikroskop.The invention relates to the series connection of variable or discrete Short pass and long pass filters to implement a freely programmable Bandpass filter for use in fluorescence microscopy, especially in a laser scanning microscope.
Diese Filter können vorteilhaft auch an neu auf den Markt kommende Farbstoffe in der Fluoreszenzfärbung angepaßt werden.These filters can also be beneficial to new dyes coming onto the market be adapted to the fluorescence staining.
In Fig. 1 ist schematisch eine Mikroskopeinheit M und ein angekoppelter Scankopf S dargestellt. Im Mikroskop ist eine Lichtquelle LQ1 mit einer Beleuchtungsoptik vorgesehen, die über einen Strahlteiler ST1 das Objekt auf dem Mikroskoptisch T konventionell beleuchtet.In Fig. 1, a microscope unit M and a coupled scan head S is shown schematically. A light source LQ1 with illumination optics is provided in the microscope, which conventionally illuminates the object on the microscope stage T via a beam splitter ST1.
Ein schwenkbarer Spiegel S3 dient zur Umschaltung auf Durchlichtbeleuchtung mittels einer Lichtquelle LQ2 über den Kondensor KO. Über eine Tubuslinse TL und einen Spiegel S1 erfolgt die Beobachtung über ein Okular OK.A swiveling mirror S3 is used to switch to transmitted light illumination by means of a light source LQ2 via the condenser KO. The observation takes place via a tube lens TL and a mirror S1 Eyepiece OK.
Über diesen Spiegel oder Strahlteiler S1 wird weiterhin der Scanstrahlengang eingekoppelt, über die Scanlinse SL und den Scanner SC.The scanning beam path continues via this mirror or beam splitter S1 coupled, via the scan lens SL and the scanner SC.
Das Licht eines Lasers L wird über Verschluß V, Einkoppeloptik EO, Spiegel S2 und Strahlteiler ST2 in Richtung des Scanners SC eingekoppelt. Über den dichroitische Strahlteiler ST2 gelangt das vom Objekt kommende Reflexions- und/oder Fluoreszenzlicht über eine Kollimationsoptik KO und ein Pinhole PH auf einen Detektor dem eine erfindungsgemäße Filtereinheit FE vorgeordnet ist, die in Fig. 1a schematisch vergrößert dargestellt ist. The light from a laser L is coupled in in the direction of the scanner SC via shutter V, coupling optics EO, mirror S2 and beam splitter ST2. Via the dichroic beam splitter ST2, the reflection and / or fluorescent light coming from the object passes through a collimating lens KO and a pinhole PH to a detector which is preceded by a filter unit FE according to the invention, which is shown schematically enlarged in FIG. 1a.
Die Filtereinheit besteht vorteilhaft aus auswechselbaren Kurzpaß- und Langpaßfiltern, wodurch sich die Breite und der Schwerpunkt eines gewünschten Bandpasses einstellen lassen. Zusätzlich kann noch ein sogenannter "Notch"-Filter vorgesehen sein, der nur die Anregungswellenlänge mit hoher optischer Dichte unterdrückt, was die Nachweisempfindlichkeit für das Fluoreszenzlicht noch erhöht. Es können natürlich auch in mehreren Detektionskanälen die beschriebenen Filtereinheiten verwendet werden, wobei die Aufteilung der Detektionskanäle in bekannter Weise, beispielsweise gemäß DE 19 70 275A1, mittels dichroitischer Strahlteiler, auch auswechselbar auf einem Teilerrevolver, zur groben Vorselektion des Spektralbereiches erfolgen kann und die Feineinstellung über die erfindungsgemäßen Filtereinheiten erfolgt.The filter unit advantageously consists of exchangeable short-pass and Long pass filters, which changes the width and center of gravity of a desired one Have bandpasses set. In addition, a so-called "notch" filter can be provided, which only the Excitation wavelength with high optical density suppresses what the Detection sensitivity for the fluorescent light still increased. Of course, the described ones can also be used in several detection channels Filter units are used, the division of the detection channels into known manner, for example according to DE 19 70 275A1, using dichroic beam splitters, also exchangeable on a divider turret, for rough preselection of the spectral range can be done and the fine adjustment takes place via the filter units according to the invention.
Fig. 2a-d zeigen Beispiele von erfindungsgemäßen Filteranordnungen, Fig. 2a hintereinandergeschaltete Filterschieber FS1, FS2 mit kreisförmigen Filtern F1 bzw. F2, wobei in optischen Achse A jeweils zwei unterschiedliche Filter miteinander kombiniert sind. 2a-d show. Examples of inventive filter arrangement, Fig. 2a cascaded filter slide FS1, FS2 with circular filters F1 and F2, respectively, two different filters are combined with each other in the optical axis A, respectively.
In Fig. 2b sind die Filter F1, F2 auf drehbaren Filterrädern FR1, FR2 angeordnet und miteinander in der optischen Achse A kombinierbar.In Fig. 2b, the filter F1, F2, and arranged on rotating filter wheels FR1 FR2 each other in the optical axis A can be combined.
Besonders vorteilhaft ist die Kombination mindestens zweier kontinuierlicher Verlaufs-Kantenfilter VF1, VF2, in Fig. 2c kreisförmig und jeweils verdrehbar in Fig. 2d verschiebbar dargestellt.The combination of at least two continuous profile edge filters VF1, VF2 is shown in a particularly advantageous manner, circular in FIG. 2c and displaceable in each case in a rotatable manner in FIG. 2d.
Derartige Kantenfilter sind beispielsweise von COHERENT (Variable Edge-pass-Filters) erhältlich.Edge filters of this type are, for example, from COHERENT (variable Edge pass filters) available.
In Fig. 2d sind die Verlaufskurven einzelner Filterabschnitte dargestellt, in VF2 als Kurzpassfilter SP, in VF1 als Langpaßfilter LP.In Fig. 2d the trajectories of individual filter sections are shown in VF2 as a short pass filter SP, in VF1 as long pass filter LP.
In Fig. 3 ist anhand von zwei unterschiedlichen Einstellungen dargestellt, wie durch Kombination der Kurz- und Langpaßfilter und gegenseitige Verdrehung beliebig breite Bandpaßfilter für die zu detektierende Wellenlänge erzeugt werden kann.In Fig. 3 is illustrated by way of two different settings as may be generated by combining the short and long pass filter and band-pass filter for mutual rotation of any width to be detected wavelength.
Besonders vorteilhaft besteht hierdurch die Möglichkeit, komplette Fluoreszenzspektren von untersuchten exogenen oder endogenen Fluophoren in der Probe durch kontinuierliches schmalbandiges Durchscannen der Wellenlänge, im Sinne eines in der Wellenlänge variablen Monochromators, aufzunehmen. Dadurch kann durch Ansteuerung des Scanners, des Probentisches und der Detektion eine wellenlängenabhängige und zeitabhängige Detektion in x,y und z-Richtung erfolgen, wodurch die fünfdimensionale Generierung (x, y, z, t, λ) eines Probenbildes erfolgt, d. h. die Aufnahme eines Spektrums zu jedem Pixel und jederzeit im Takt der jeweiligen Abtastung.This makes it particularly advantageous for complete Fluorescence spectra of examined exogenous or endogenous fluophores in the sample by continuously scanning the wavelength, in the sense of a variable wavelength monochromator. This can be done by controlling the scanner, the sample table and the Detection a wavelength-dependent and time-dependent detection in x, y and z direction, whereby the five-dimensional generation (x, y, z, t, λ) a sample picture is taken, d. H. recording a spectrum for each pixel and at any time in time with the respective scan.
Die erfindungsgemäßen Filtereinheiten können weiterhin vorteilhaft zur spektralen Selektion auf der Beleuchtungsseite eingesetzt werden, beispielsweise an der Stelle des Verschlusses V, beispielsweise zur spektralen Einengung einer breitbandigen Laserlichtquelle oder zur Wellenlängenselektion bei einer HBO oder HAL-Lampe.The filter units according to the invention can furthermore advantageously be spectral Selection can be used on the lighting side, for example on the spot of the closure V, for example for the spectral narrowing of a broadband Laser light source or for wavelength selection with an HBO or HAL lamp.
Claims (8)
Priority Applications (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
DE1998135070 DE19835070B4 (en) | 1998-08-04 | 1998-08-04 | Arrangement for adjustable wavelength-dependent detection in a fluorescence microscope |
JP21781199A JP4608688B2 (en) | 1998-08-04 | 1999-07-30 | Laser scanning fluorescence microscope and image recording method |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
DE1998135070 DE19835070B4 (en) | 1998-08-04 | 1998-08-04 | Arrangement for adjustable wavelength-dependent detection in a fluorescence microscope |
Publications (2)
Publication Number | Publication Date |
---|---|
DE19835070A1 true DE19835070A1 (en) | 2000-02-10 |
DE19835070B4 DE19835070B4 (en) | 2006-03-16 |
Family
ID=7876336
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
DE1998135070 Revoked DE19835070B4 (en) | 1998-08-04 | 1998-08-04 | Arrangement for adjustable wavelength-dependent detection in a fluorescence microscope |
Country Status (2)
Country | Link |
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JP (1) | JP4608688B2 (en) |
DE (1) | DE19835070B4 (en) |
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DE10148782A1 (en) * | 2001-09-28 | 2003-04-24 | Zeiss Carl Jena Gmbh | Optical modulation element |
DE10156695B4 (en) * | 2001-11-17 | 2004-07-15 | Leica Microsystems Heidelberg Gmbh | Scanning microscope, scanning microscopy method and bandpass filter |
US7075716B2 (en) | 2000-07-17 | 2006-07-11 | Leica Microsystems Cms Gmbh | Arrangement for spectrally sensitive reflected-light and transmitted-light microscopy |
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US11988605B2 (en) | 2018-10-22 | 2024-05-21 | Robert Bosch Gmbh | Filter device for an optical module for a lab-on-a-chip analysis device, optical module for a lab-on-a-chip analysis device and method for operating an optical module for a lab-on-a-chip analysis device |
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US7075716B2 (en) | 2000-07-17 | 2006-07-11 | Leica Microsystems Cms Gmbh | Arrangement for spectrally sensitive reflected-light and transmitted-light microscopy |
DE10148782A1 (en) * | 2001-09-28 | 2003-04-24 | Zeiss Carl Jena Gmbh | Optical modulation element |
DE10156695B4 (en) * | 2001-11-17 | 2004-07-15 | Leica Microsystems Heidelberg Gmbh | Scanning microscope, scanning microscopy method and bandpass filter |
US6954306B2 (en) | 2001-11-17 | 2005-10-11 | Leica Microsystems Heidelberg Gmbh | Scanning microscope, method for scanning microscopy, and bandpass filter |
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Also Published As
Publication number | Publication date |
---|---|
JP2000056228A (en) | 2000-02-25 |
DE19835070B4 (en) | 2006-03-16 |
JP4608688B2 (en) | 2011-01-12 |
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