U.S. patent application number 09/988880 was filed with the patent office on 2002-06-06 for rapidly changing dichroic beamsplitter.
Invention is credited to Stuckey, Jeffrey A..
Application Number | 20020068020 09/988880 |
Document ID | / |
Family ID | 22944311 |
Filed Date | 2002-06-06 |
United States Patent
Application |
20020068020 |
Kind Code |
A1 |
Stuckey, Jeffrey A. |
June 6, 2002 |
Rapidly changing dichroic beamsplitter
Abstract
An epi-fluorescent microscope is constructed with a quick
changing set of filters. A wheel containing a plurality of dichroic
beamsplitter mirrors mounted equidistance from a center axis of the
wheel is mounted at a 45.degree. angle within the lightpath and
provides a beamsplitter function. Light from an excitation source
is provided as monochromatic light, so that light from the
excitation source is reflected by a selected one of the dichroic
mirrors to the specimen, and Stokes shifted light from the specimen
is transmitted through the dichroic mirror. By separately mounting
the dichroic mirrors, the dichroic mirrors can be quickly moved and
a quicker change of sensed fluorophores is achieved.
Inventors: |
Stuckey, Jeffrey A.;
(Jeffersonville, PA) |
Correspondence
Address: |
ELMAN & ASSOCIATES
P O BOX 1969
MEDIA
PA
19063-8969
US
|
Family ID: |
22944311 |
Appl. No.: |
09/988880 |
Filed: |
November 19, 2001 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
60249626 |
Nov 17, 2000 |
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Current U.S.
Class: |
422/82.08 ;
250/458.1; 359/634 |
Current CPC
Class: |
G02B 21/06 20130101;
G02B 26/007 20130101; G02B 21/16 20130101 |
Class at
Publication: |
422/82.08 ;
359/634; 250/458.1 |
International
Class: |
G01N 021/64 |
Foreign Application Data
Date |
Code |
Application Number |
Nov 16, 2001 |
US |
PCT/US01/42957 |
Claims
1. A microscope with apparatus for rapidly changing dichroic
beamsplitters in epi_fluorescent microscopes, the microscope
comprising: a source of excitation light; a high speed wheel having
a center axis bisecting a predetermined reflection angle within an
epi-fluorescent lightpath of the microscope optics and rotable
about the center axis driven by a changer; a plurality of dichroic
beamsplitter mirrors mounted to the wheel; the wheel mounted in an
epi-fluorescent microscope; and a computer control, the changer
responsive to the changer computer control, wherein computer
software can command the changer to rotate different ones of the
plurality of dichroic beamsplitters into the epi_fluorescent
lightpath to position a selected one of the dichroic beamsplitter
mirrors in the lightpath to image a particular fluorophore, thereby
providing a microscope system in which the device is commanded by
the software to change dichroic beamsplitters.
Description
CROSS REFERENCE TO RELATED APPLICATIONS
[0001] This application claims priority from U.S. Provisional
Patent Application No. 60/249,626, filed Nov. 17, 2000, and is a
continuation International Patent Application PCT/US01/42957. The
entire contents of both of the aforementioned patent applications
are incorporated herein by reference.
BACKGROUND OF THE INVENTION
[0002] This invention relates to epi-fluorescent microscopy, and
more particularly to the rapid selection of filters for measuring
fluorescence at different wavelengths or Stokes shifts.
[0003] Current fluorescent microscope designs employ an incident
light or epi-fluorescent design where a dichroic beam splitter (or
chromatic mirror) mounted in a filter cube at a 45 degree angle to
the excitation light path, is used to reflect shorter excitation
wavelengths of light onto the specimen while passing longer
emission wavelengths to the eyepieces or camera (FIG. 2).
[0004] Many fluorescence applications require two or more
fluorescent labels to be present in the specimen. Each label has
its own excitation and emission spectra, and thus requires
different excitation and emission filters, as well as a different
dichroic beamsplitter.
[0005] To date two approaches to using multiple fluorophores have
been employed:
[0006] 1) Use dichroic beamsplitters that have multiple cutoff
wavelengths. Thus a single dichroic can be used with multiple
fluorophores. However because of bandwidth restrictions, total
light throughput is reduced, thus creating longer exposure times
when working with a camera. Longer exposure times translate into
longer acquisition duty cycles which is problematic in paradigms
requiring repetitive high-speed data acquisition (screening
applications and applications using living cells).
[0007] 2) Motorized filter cube changers. Several commercially
available microscopes employ motorized filter cube changes. These
allow the use of single dichroics for each fluorophore. However
switching time is slow (1-2 seconds) which creates problems when
using multiple fluorophores in paradigms requiring repetitive
high-speed data acquisition (screening applications and
applications using living cells).
SUMMARY OF INVENTION
[0008] Disclosed below is a device for rapidly changing dichroic
beamsplitters in epi-fluorescent microscopes. The device is a high
speed wheel in which dichroic beamsplitters are mounted. The high
speed dichroic changer is mounted in an epi-fluorescent microscope,
and the changer is under computer control. Computer software can
command the changer to rotate different dichroic beamsplitters into
the epifluorescent lightpath so that the appropriate dichroic is in
position when a particular fluorophore is imaged. The present
invention provides a microscope system in which the device is
commanded by the software to change dichroic beamsplitters (FIG.
1).
BRIEF DESCRIPTION OF THE DRAWINGS
[0009] FIG. 1 is a system schematic showing some key components of
an analysis microscope constructed according to the present
invention.
[0010] FIG. 2 is a schematic drawing showing epi-fluorescent light
paths occurring in the system of FIG. 1.
[0011] FIG. 3 is a top view showing the arrangement of a high speed
dichroic beamsplitter wheel constructed according the present
invention.
[0012] FIG. 4 is a schematic drawing showing control of a
microscope according to one aspect of the invention.
DETAILED DESCRIPTION
[0013] FIG. 1 is a system schematic showing some key components of
an analysis microscope 11 constructed according to one aspect of
the invention. The microscope includes a light source 12, a
dichroic beamsplitter wheel 13, and an objective 14 which is
aligned with a specimen 15. light from the light source 12 is
reflected at the dichroic wheel 13, and passes through the
objective 14. Light reflected or fluorescing from the specimen 15
again passes through the objective 14, and that light which passes
through the dichroic wheel 13 is received by the camera 16. Still
referring to FIG. 1, the dichroic wheel 13 is rotationally
positioned by a stepper motor 17, which is controlled by motor
controller 18. The motor controller 18 is responsive to a computer
19, and the computer 19 also receives images from the camera
16.
[0014] FIG. 2 is a schematic drawing showing epi-fluorescent light
paths. Light from the light source 12 is reflected by the dichroic
beamsplitter wheel 13 which is at a reflection angle which
determines the lightpath. In the exemplary embodiment, a 45 degree
angle is used as the reflection angle, although since standard
dichroic cubes with the 45 degree angle are not used, it is likely
that other reflection angles can be used. The return light, if it
is able to pass the dichroic wheel 13, is received by the camera.
This sequence generally corresponds to fluorescent microscope
designs which employ a dichroic beam splitter or a chromatic mirror
mounted in a filter cube at a 45 degree angle to the excitation
light path. Significantly the filter cube is not required.
[0015] FIG. 3 is a top view showing the arrangement of a high speed
dichroic beamsplitter wheel 13 constructed according to one
embodiment of the invention. The wheel 13 includes a support plate
52, on which a plurality of dichroic beamsplitter lenses 61-65 are
arranged about a center axis 66 of the support plate 52. Each of
the lenses 61-65 have distinct filtering properties. In many cases,
only two lenses are required, so it is possible to use plano lenses
in the remaining three spots, or to leave the remaining three spots
empty.
[0016] The solution described here is to mount round 50 mm dichroic
beamsplitters in a high-speed filter wheel (FIG. 3). Any size or
shape dichroic beamsplitter that matched the optical path of the
microscope or instrument would be acceptable.
[0017] The wheel is capable of switching between adjacent dichroics
in 50 msec. The wheel in the current implementation is driven by a
DC stepper motor and is under computer control. Switching time is a
function of wheel mass and motor speed. Any motor-wheel combination
that allowed faster switching times would be acceptable. It is also
conceivable that a galvanometer could drive a wheel at much higher
speeds, or that a galvanometer could also move dichroic
beamsplitters mounted on a spindle.
[0018] The wheel is mounted at a 45 degree angle inside the
microscope such that when a given dichroic is selected the opening
holding that dichroic is rotated into the epi-fluorescent light
path, and the selected dichroic beamsplitter is at 45 degree angle
to the excitation light (FIG. 1, FIG. 4).
[0019] This design this allows optimized throughput for each
fluorophore while at the same time being able to switch the
dichroics rapidly. Optimized throughput is advantageous as it
allows for shorter camera exposure times, which decreases duty
cycle as well as helps reduce photobleaching. Being able to rapidly
switch the dichroic beamsplitter reduces the interval between image
acquisitions, thus reducing duty cycle.
[0020] FIG. 4 is a schematic drawing showing control for a
microscope using a fast dichroic beamsplitter changer according to
one aspect of the invention. The figure depicts a loop 80, which
represents one complete duty cycle with respect to a predetermined
wavelength. Application software sends a move command 82 to a
dichroic beamsplitter wheel controller such as controller 18 in
FIG. 1. The controller firmware then interprets 83 the move command
and rotates 83 the wheel (13, FIG. 1) to a correct position. The
controller 18 then signals 84 application software that wheel 13
has finished moving. In response, the computer (19, FIG. 1) signals
85 the camera (16, FIG. 1) to acquire 86 an image. The image is
then acquired 87.
[0021] This sequence repeated by executing a loop 80 for every
wavelength required.
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