U.S. patent application number 11/734208 was filed with the patent office on 2007-08-09 for total reflection fluorescent microscope.
This patent application is currently assigned to OLYMPUS CORPORATION. Invention is credited to Yasushi Aono, Kenichi Kusaka, Atsuhiro TSUCHIYA.
Application Number | 20070183030 11/734208 |
Document ID | / |
Family ID | 33487104 |
Filed Date | 2007-08-09 |
United States Patent
Application |
20070183030 |
Kind Code |
A1 |
TSUCHIYA; Atsuhiro ; et
al. |
August 9, 2007 |
TOTAL REFLECTION FLUORESCENT MICROSCOPE
Abstract
A fluorescent microscope comprises a light source, an optical
illumination system which forms an optical path to irradiate a
specimen with a light beam from the light source, an objective lens
which condenses the light beam of the optical illumination system
onto the specimen, an optical device which is disposed on the
optical path of the optical illumination system and which decenters
the light beam by decentering an optical axis of the optical path,
and a slit which passes the light beam decentered by the optical
device through a total reflection illumination region on an
emission pupil surface of the objective lens.
Inventors: |
TSUCHIYA; Atsuhiro;
(Hachioji-shi, JP) ; Kusaka; Kenichi;
(Akiruno-shi, JP) ; Aono; Yasushi; (Yokohama-shi,
JP) |
Correspondence
Address: |
FRISHAUF, HOLTZ, GOODMAN & CHICK, PC
220 Fifth Avenue
16TH Floor
NEW YORK
NY
10001-7708
US
|
Assignee: |
OLYMPUS CORPORATION
Tokyo
JP
|
Family ID: |
33487104 |
Appl. No.: |
11/734208 |
Filed: |
April 11, 2007 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
10848626 |
May 18, 2004 |
7224524 |
|
|
11734208 |
Apr 11, 2007 |
|
|
|
Current U.S.
Class: |
359/388 |
Current CPC
Class: |
G02B 21/16 20130101;
G02B 21/10 20130101; G02B 21/0088 20130101 |
Class at
Publication: |
359/388 |
International
Class: |
G02B 21/06 20060101
G02B021/06 |
Foreign Application Data
Date |
Code |
Application Number |
May 21, 2003 |
JP |
2003-143382 |
Claims
1. A fluorescent microscope comprising: a light source; an optical
illumination system which forms an optical path to irradiate a
specimen with a light beam from the light source; an objective lens
which condenses the light beam from the light source, which has
traveled through the optical illumination system, onto the
specimen; and a slit which is located in the optical illumination
system to pass the light beam from the light source through a total
reflection illumination region on an emission pupil surface of the
objective lens, wherein an emission position of the light beam
emitted from the light source is movable between an optical axis of
the optical illumination system and a position shifted from the
optical axis by a predetermined distance.
2. The fluorescent microscope according to claim 1, wherein the
optical illumination system comprises: a collector lens which
parallelizes the light beam from the light source; a condenser
which condenses the light beam from the light source that has
passed through the collector lens; and a projection lens which
projects an image formed by the light beam condensed by the
condenser onto an emission pupil surface of the objective lens.
3. The fluorescent microscope according to claim 1, wherein the
slit is removed from the optical illumination system, and the
emission position of the light beam emitted from the light source
is moved onto the optical axis of the optical illumination system,
to perform fluorescent illumination.
4. A fluorescent microscope comprising: an objective lens which is
used in observation of a specimen; a movable light source which
irradiates the specimen with a light beam; a collector lens which
parallelizes the light beam from the light source; a condenser
which condenses the light beam from the light source that has
passed through the collector lens; and a slit which is located at a
position onto which the light beam is condensed by the condenser; a
projection lens which projects an image formed by the light beam
from the light source through the slit onto an emission pupil
surface of the objective lens, wherein when the light source is
moved to a position displaced at least from an optical axis of the
collector lens, the image formed by the light beam from the light
source is projected onto the slit, and the light beam from the
light source is caused to pass through a total reflection
illumination region of the emission pupil surface of the objective
lens.
5. The fluorescent microscope according to claim 4, wherein the
light source is moved onto the optical axis of the collector lens
and the slit is removed from a path of the light beam from the
light source to perform fluorescent illumination.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] The present application is a Divisional Application of U.S.
application Ser. No. 10/848,626 filed May 18, 2004, which is based
upon and claims the benefit of priority from prior Japanese Patent
Application No. 2003-143382, filed May 21, 2003, the entire
contents of which are incorporated herein by reference.
BACKGROUND OF THE INVENTION
[0002] 1. Field of the Invention
[0003] The present invention relates to a total reflection
fluorescent microscope, which can perform fluorescent observation
by total reflection illumination.
[0004] 2. Description of the Related Art
[0005] In recent years, functions of biological cells have been
vigorously analyzed. In the function analysis of the cells,
attentions have been paid especially to a total reflection
fluorescent microscope which acquires a total reflection
fluorescent image from a cell membrane and its vicinity as a device
for observing the function of the cell membrane.
[0006] Total reflection illumination which locally illuminates only
a sample in the vicinity of a glass surface is used in the total
reflection fluorescent microscope. In the total reflection
illumination, an evanescent light is used which oozes to a sample
side by about several hundreds of nanometers in a boundary surface
between glass and sample, and a background noise (scattered light
and the like) is remarkably low. Therefore, fluorescent observation
of even a molecular of fluorescent dyestuff is possible by the
total reflection fluorescent microscope.
[0007] Additionally, in general, in the total reflection
fluorescent microscope, a laser light beam is used as a light
source. A total reflection fluorescent microscope in which the
laser light beam is introduced into an optical illumination system
of the microscope via a glass fiber is described, for example, in
Jpn. Pat. Appln. KOKAI Publication No. 2002-169097.
[0008] However, the laser light source which produces the laser
light beam is expensive, and additionally a monochromatic light is
produced. Therefore, for example, in order to cope with fluorescent
reagents having various excitation wavelength characteristics, a
plurality of laser light sources have to be prepared. Therefore,
the total reflection fluorescent microscope becomes further
expensive, and additionally a large occupying space is also
required for installing a plurality of laser light sources.
[0009] To solve the problem, as described in Jpn. Pat. Appln. KOKAI
Publication No. 2002-236258, a total reflection fluorescent
microscope has been proposed in which inexpensive white light
sources such as a mercury lamp and a xenon lamp are used instead of
the laser light source. The total reflection fluorescent microscope
according to the Jpn. Pat. Appln. KOKAI Publication No. 2002-236258
is configured as follows. A ring slit for transmitting a light beam
in an annular form is disposed in the optical illumination system
disposed on an optical path of the light emitted from the white
laser light beam. Moreover, when an image of the ring slit is
projected onto an emission pupil surface of an objective lens, an
illuminative light is guided only to an orbicular total reflection
region around an emission pupil of the objective lens. Moreover,
total reflection is performed in a boundary surface between a
specimen and cover glass to produce is the evanescent light, and a
fluorescent dyestuff is excited.
BRIEF SUMMARY OF THE INVENTION
[0010] A fluorescence microscope according to a first aspect of the
present invention is characterized by comprising a light source; an
optical illumination system which forms an optical path to
irradiate a specimen with a light beam from the light source; an
objective lens which condenses the light beam of the optical
illumination system onto the specimen; an optical device which is
disposed on the optical path of the optical illumination system and
which decenters the light beam by decentering an optical axis of
the optical path; and a slit which passes the light beam decentered
by the optical device through a total reflection illumination
region on an emission pupil surface of the objective lens.
[0011] A fluorescence microscope according to a second aspect of
the present invention is characterized by comprising: a light
source; an optical illumination system which forms an optical path
to irradiate a specimen with a light beam from the light source; an
objective lens which condenses the light beam of the optical
illumination system onto the specimen; and a slit which passes the
light beam from the light source through a total reflection
illumination region on an emission pupil surface of the objective
lens, in which an emission position of the light beam emitted from
the light source is movable between an optical axis of the optical
illumination system and a position shifting from the optical axis
by a predetermined distance.
[0012] Advantages of the invention will be set forth in the
description which follows, and in part will be obvious from the
description, or may be learned by practice of the invention. The
objects and advantages of the invention may be realized and
obtained by means of the instrumentalities and combinations
particularly pointed out hereinafter.
BRIEF DESCRIPTION OF THE SEVERAL VIEWS OF THE DRAWINGS
[0013] The accompanying drawings, which are incorporated in and
configure a part of the specification, illustrate embodiments of
the invention, and together with the general description given
above and the detailed description of the embodiments given below,
serve to explain the principles of the invention.
[0014] FIG. 1 is a diagram showing a schematic configuration of a
first embodiment of the present invention;
[0015] FIG. 2 is a diagram showing a schematic configuration of a
main part of the first embodiment;
[0016] FIGS. 3A to 3C are explanatory views of a slit for use in
the first embodiment;
[0017] FIG. 4 is an explanatory view of a state in which a wedge
prism and slit are removed from an optical path is of an optical
illumination system in the first embodiment;
[0018] FIG. 5 is a diagram showing a schematic configuration of a
main part of a second embodiment;
[0019] FIG. 6 is a diagram showing a light flux refracted by the
wedge prism for use in the second embodiment:
[0020] FIGS. 7A to 7D are explanatory views of a slit having a
crescent opening for use in the second embodiment;
[0021] FIGS. 8A to 8E are explanatory views of a slit having a
small-diameter opening for use in the second embodiment;
[0022] FIGS. 9A to 9D are explanatory views of a slit having an
annular opening for use in the second embodiment;
[0023] FIGS. 10A and 10B are diagrams showing a schematic
configuration of a main part of a modification of the second
embodiment;
[0024] FIG. 11 is a diagram showing a schematic configuration of a
third embodiment of the present invention;
[0025] FIG. 12 is a diagram showing a light flux refracted by a
conical prism for use in the third embodiment;
[0026] FIG. 13 is an explanatory view of a state of a light source
image projected on the annular opening of the third embodiment;
[0027] FIGS. 14A and 14B are diagrams showing a schematic
configuration of a main part of a modification of the third
embodiment;
[0028] FIG. 15 is a diagram showing a schematic configuration of a
fourth embodiment of the present invention;
[0029] FIG. 16 is an explanatory view of an LED image projected on
the crescent opening of the fourth embodiment;
[0030] FIG. 17 is an explanatory view of a state of an LED image
projected on the annular opening of the fourth embodiment;
[0031] FIG. 18 is a diagram showing a schematic configuration of a
fifth embodiment of the present invention;
[0032] FIGS. 19A and 19B are explanatory views of a slit having a
crescent opening for use in the fifth embodiment;
[0033] FIGS. 20A and 20B are explanatory views of a slit having an
annular opening for use in the fifth embodiment;
[0034] FIG. 21 is an explanatory view of a slit showing a
small-diameter opening for use in the fifth embodiment;
[0035] FIG. 22 is an explanatory view of a state in which an afocal
converter, wedge prism, and slit are detached from an optical path
of an optical illumination system in the fifth embodiment;
[0036] FIG. 23 is a diagram showing a schematic configuration of a
sixth embodiment of the present invention;
[0037] FIG. 24 is a diagram showing a schematic configuration of a
seventh embodiment of the present invention; and
[0038] FIG. 25 is a diagram showing a schematic configuration of an
eighth embodiment of the present invention.
DETAILED DESCRIPTION OF THE INVENTION
[0039] Embodiments of the present invention will be described
hereinafter with reference to the drawings.
FIRST EMBODIMENT
[0040] FIG. 1 is a diagram showing a schematic configuration of a
total reflection fluorescent microscope to which the present
invention is applied. In this case, FIG. 1 shows an example of an
inverted microscope for performing observation by an objective lens
disposed below a specimen.
[0041] In FIG. 1, a stage 2 is disposed in an upper part of a
microscope main body 1. A specimen 3 is laid on the stage 2. In
this case, as shown in FIG. 2, a cover glass 7 is disposed under
the specimen 3. An objective lens 4 is disposed under the cover
glass 7 via oil (not shown).
[0042] A revolver 5 is disposed under the specimen 3. The revolver
5 is held in the microscope main body 1. The revolver 5 holds a
plurality of objective lenses 4. When the revolver 5 rotates, it is
possible to selectively dispose the objective lens 4 having a
magnification or a type required for the observation on an
observation optical axis 6. When the revolver 5 vertically moves
along the observation optical axis 6 by an operation of a focusing
handle (not shown) to change a relative distance between the
specimen 3 and the objective lens 4 on the stage 2, the specimen 3
can be focused.
[0043] A light source 11 for illuminating the specimen 3 is used in
total reflection fluorescent illumination or usual fluorescent
illumination in which total reflection is not performed. As the
light source 11, high-luminance arc light sources such as a mercury
lamp and xenon lamp are used. It is to be noted that these arc
light sources preferably have micro luminescent spots, and the
light source is selected having a luminescent spot in which a
projected image on an emission pupil surface of the objective lens
is smaller than an emission pupil diameter of the objective
lens.
[0044] A collector lens 12 is provided on an illuminative light
axis 18 of the optical path from the light source 11. The collector
lens 12 condenses the light beams from the light source 11 and
emits a parallel light beam.
[0045] A wedge-shaped plane plate 41 for decentering the optical
path is disposed in the optical path of the parallel light beam
from the collector lens 12. The wedge-shaped plane plate 41
refracts the parallel light beam emitted from the collector lens 12
at a predetermined angle with respect to the illuminative light
axis 18.
[0046] A condenser 14 and a slit 15 are disposed in the optical
path of a light flux refracted by the wedge-shaped plane plate 41.
The condenser 14 condenses the light flux refracted by the
wedge-shaped plane plate 41 on a surface of a slit 15, and a light
source image 11a of the light source 11 is projected.
[0047] In the slit 15, slits having openings 43a, 43b, 43c having
three different shapes shown in FIGS. 3A to 3C are used. These
openings 43a, 43b, 43c transmit the light beams (light source
images 11a) condensed by the condenser 14. The light beams
transmitted through the openings 43a, 43b, 43c are to be totally
reflected by a boundary surface between the specimen 3 and the
cover glass 7.
[0048] It is to be noted that in any of three types of slits 15,
two slits are disposed closely along an illuminative light axis 18.
This is because the light beam that is not totally reflected,
generated by frame reflection inside the illuminative light axis or
by a diffracted light in the slit 15, is cut. The wedge-shaped
plane plate 41 and slit 15 move along a plane vertical to the
illuminative light axis 18 by known switching mechanism such as a
slider, and are detachably inserted with respect to the optical
path. When the wedge-shaped plane plate 41 and slit 15 are inserted
into the optical path, the total reflection fluorescent
illumination can be selected. When the wedge-shaped plane plate 41
and slit 15 are removed from the optical path, the usual
fluorescent illumination that does not perform the total reflection
can be selected. The slit 15 is movable along a plane crossing the
illuminative light axis 18 at right angles (in a vertical direction
of an arrow shown in FIG. 1 in this case) in a state in which the
slit is inserted in the optical path.
[0049] A field stop (FS) 16 and an FS projection lens 17 are
disposed in the optical path of the light transmitted through the
slit 15. The field stop (FS) 16 is used to restrict an illumination
field, and a slit diameter can be varied. The FS projection lens 17
projects the field stop (FS) 16 on three surfaces of the specimen
3, and projects the image of the slit 15 on an emission pupil
surface of the objective lens 4.
[0050] A rotatable cassette 19 which holds two or more mirror units
18a, 18b is disposed on the optical path where the light beam is
transmitted through the FS projection lens 17. The mirror units
18a, 18b are detachably fixed to the cassette 19 by known means
such as a dovetail.
[0051] The cassette 19 is rotated around a rotation axis 20. By
this rotation, the mirror units 18a, 18b are selectively switched
on the observation optical axis 6 in accordance with wavelength
characteristics of a fluorescent dyestuff with which the specimen 3
is dyed. In FIG. 1, the mirror unit 18a is switched (disposed) on
the observation optical axis 6.
[0052] In the mirror unit 18a, an excitation filter 211, dichroic
mirror 212, and absorption filter 213 are integrally disposed as a
set. The excitation filter 211 selectively transmits a wavelength
required for exciting the specimen 3 among the light beams emitted
from the FS projection lens 17. The dichroic mirror 212 reflects an
excitation wavelength from the excitation filter 211, and transmits
a fluorescent wavelength emitted from the specimen 3. The dichroic
mirror 212 is inclined by 45.degree. with respect to both the
illuminative light axis 18 and the observation optical axis 6 in
such a manner that an excited light along the illuminative light
axis 18 from the excitation filter 211 is guided in a direction
matching the optical axis (observation optical axis 6). The
absorption filter 213 selectively transmits only the wavelength
required for the observation in fluorescence emitted from the
specimen 3.
[0053] An optical relay system 231 is disposed in a transmission
optical path of the absorption filter 213. An image of the specimen
3 formed by the objective lens 4 is relayed to the vicinity of an
eyepiece lens 232. The eyepiece lens 232 is used in such a manner
that the image of the specimen 3 relayed through the optical relay
system 231 is visually observable.
[0054] Since the mirror unit 18b is configured in the same manner
as in the mirror unit 18a, the description is omitted.
[0055] A transmission illumination section 26 includes an optical
transmission illumination system in a case where transmission
illumination observation is performed.
[0056] A case where the total reflection fluorescent illumination
is performed in the above-described configuration will be
described.
[0057] When an illuminative light beam is emitted from the light
source 11, the light beam is formed into the parallel light beam by
the collector lens 12 and is incident upon the wedge-shaped plane
plate 41.
[0058] The wedge-shaped plane plate 41 refracts the parallel light
beam from the collector lens 12 at a predetermined angle to emit a
light flux (light source image) refracted with respect to the
illuminative light axis 18. The light flux is condensed onto the
slit 15 by the condenser 14.
[0059] In this case, in the slit 15, the crescent opening 43a (or
the opening 43b or 43c) is formed linearly symmetrically with
respect to a line passing through a center of the slit 15 on a slit
15 surface as shown in FIG. 3A. A circular portion of the opening
43a is formed substantially in parallel with a circumferential
direction (i.e., on a substantially concentric circle).
[0060] Moreover, the light source image 11a is projected as the
image of the light source 11 in the crescent opening 43a of the
slit 15.
[0061] In the light source image 11a projected on the slit 15, the
light beam transmitted through the opening 43a is incident upon the
excitation filter 211 via the FS projection lens 17. The excitation
filter 211 selects the light beam having a wavelength required for
exciting the specimen 3. The light beam selected by its wavelength
is reflected toward the objective lens 4 by the dichroic mirror
212, and subsequently projected onto the emission pupil of the
objective lens 4. It is to be noted that the image of the slit 15
projected on the emission pupil surface of the objective lens 4,
that is, a slit image will be described later.
[0062] All the light beams transmitted through the opening 43a of
the slit 15 form totally reflected light beams within a total
reflection region.
[0063] As shown in FIG. 2, the light beam transmitted through the
emission pupil surface of the objective lens 4 passes through a
peripheral edge portion of the objective lens 4, and reaches the
cover glass 7 via the oil charged between the objective lens 4 and
the cover glass 7. Here, the total reflection occurs in the
boundary surface between the specimen 3 and the cover glass 7, and
the evanescent light is generated in a range of about 50 to 200 nm
on a specimen side of the boundary surface. The fluorescent
dyestuff with which the specimen 3 is dyed by the evanescent light
is excited to emit the fluorescence.
[0064] In this state, a surveyor moves the stage 2 to search for a
desired observation range on the specimen 3, vertically moves the
revolver 5 along the observation optical axis 6 by the operation of
the focusing handle (not shown), and changes the relative distance
between the specimen 3 and the objective lens 4 to focus the
specimen 3.
[0065] The fluorescence emitted from the specimen 3 passes through
the dichroic mirror 212, and the fluorescent wavelength required
for the observation is selected by the absorption filter 213.
Moreover, the image of the specimen 3 formed by the objective lens
4 is relayed to the eyepiece lens 232 via the optical relay system
231, and visual observation is possible.
[0066] To change the fluorescent wavelength to be observed of the
fluorescent dyestuff dyed on the specimen 3, the cassette 19 is
rotated around the rotation axis 20 and, for example, the mirror
unit 18b may be switched on the observation optical axis 6 instead
of the mirror unit 18a. To change the observation magnification of
the specimen 3, the revolver 5 may be rotated to position the
objective lens 4 having a desired magnification on the observation
optical axis 6.
[0067] A relation between the slit image and the emission pupil
surface of the objective lens 4 by the crescent opening 43a of the
slit 15 will be described later.
[0068] In the above-mentioned description, the crescent opening 43a
is formed in the slit 15, but the small-diameter opening 43b may
also be formed in a predetermined position on the slit 15 plane,
for example, as shown in FIG. 3B, the annular opening 43c may also
be formed along the peripheral edge portion of the slit 15 surface
as shown in FIG. 3C, or an elliptic opening (not shown) may also be
formed. A relation between the slit image by the small-diameter
opening 43b, the annular opening 43c, or the elliptic opening and
the emission pupil surface of the objective lens 4 will be
described later.
[0069] A case where the total reflection fluorescent observation is
switched to usual fluorescent observation to perform the
observation will be described.
[0070] In this case, as shown in FIG. 4, the wedge-shaped plane
plate 41 and slit 15 are removed from the optical path of the
optical illumination system, and an aperture stop (AS) 29 is
inserted instead of the slit 15. Since the slit 15 is used for
transmitting the illuminative light of the light source image
through a total reflection region 27 of the objective lens 4, the
slit 15 is removed from the optical path, and the aperture stop
(AS) 29 is inserted as a diaphragm for adjusting brightness
instead. The wedge-shaped plane plate 41 is used to project the
light source image 11a in the periphery of the emission pupil of
the objective lens 4. Especially, when the objective lens 4 has a
high magnification, and the emission pupil diameter is small, the
illuminative light is reflected by the objective lens 4, the region
is darkened, and illumination unevenness sometimes occurs.
Therefore, the wedge-shaped plane plate 41 is removed from the
optical path.
[0071] From this state, visual sample observation is possible using
a generally known usual fluorescent observation method.
[0072] As described above, since the wedge-shaped plane plate 41 is
disposed as an optical device capable of projecting the light
source image 11a on an optical path between the light source 11 and
the slit 15, the optical axis of the optical path is decentered and
moved, for example, to the opening 43a of the slit 15. Therefore,
since the illuminative light can be efficiently taken into the
total reflection region 27 of the objective lens 4 having an
emission pupil diameter, the total reflection fluorescent
observation by sufficient brightness can be realized. In the
observation in the usual fluorescent illumination, when the
wedge-shaped plane plate 41 and slit 15 are removed from the
optical path, the light source image is projected on the optical
axis. Therefore, also in this case, since the illuminative light
can be efficiently taken in, the usual fluorescent observation by
the sufficient brightness can be realized.
[0073] Moreover, the slit 15 for performing the total reflection
illumination has the sector opening 43b for transmitting the light
beam only in a part of the total reflection region. Therefore, even
when the position or the size of the slit image changes by
eccentricity of the optical illumination system or magnification
error, the slit image can be prevented from deviating from the
total reflection region of the emission pupil surface of the
objective lens 4. Therefore, the slit image does not enter the
fluorescent illumination region where the total reflection is not
performed, the eccentricity error can be prevented, and the total
reflection fluorescent observation can be realized with good
contrast.
[0074] The opening 43a is formed in a crescent shape to enlarge an
opening area of a portion which is not easily influenced by
contrast deterioration with respect to the eccentricity.
Conversely, an opening area of a portion easily influenced by the
contrast deterioration with respect to the eccentricity can be
reduced. Therefore, since the influence of the contrast
deterioration by the eccentricity error of the optical system
cannot be easily exerted, and additionally a middle portion of the
opening 43a has a maximum necessary opening area, the illuminative
light from the light source 11 can be efficiently taken. Therefore,
the total reflection fluorescent observation can be realized with a
sufficient brightness and with good contrast and balance.
[0075] The slit 15 having the small-diameter opening 43c is strong
especially against the deterioration of the contrast by the
eccentricity of the optical illumination system. When the slit is
combined with the light source 11 having sufficient luminance, it
is easy to apply the slit even to an optical system which does not
have good accuracy. Since the slit 15 is easily worked, the slit is
inexpensive. Furthermore, since the shape of the opening matches
that of the luminescent spot of a general high-luminance arc light
source, the illuminative light can be efficiently taken in, and the
total reflection fluorescent observation by the sufficient
brightness can be realized.
[0076] The annular and elliptic openings will be explained in the
second embodiment.
[0077] Furthermore, the wedge-shaped plane plate 41 is disposed
between the collector lens 12 which projects the light source 11 as
the parallel light beam and the condenser 14 which condenses the
parallel light beam to form the light source image 11a on the slit
15 plane, and each parallel light beam in a light flux 22 is
refracted by the same angle by the wedge-shaped plane plate 41.
Therefore, little aberration is caused by the wedge-shaped plane
plate 41 or the condenser 14, and the satisfactory light source
image 11a can be projected on the slit 15 plane. Accordingly, the
illuminative light can be efficiently taken in, and bright total
reflection fluorescent observation can be performed.
SECOND EMBODIMENT
[0078] Next, a second embodiment of the present invention will be
described.
[0079] FIG. 5 is a diagram showing a schematic configuration of a
main part of the second embodiment, and the same components as
those of FIG. 1 are denoted with the same reference numerals.
[0080] In the second embodiment, a wedge prism 13 is disposed as an
optical device for decentering the optical axis in the optical path
of the parallel light beam from the collector lens 12. The wedge
prism 13 refracts the parallel light beam emitted from the
collector lens 12 in two directions including upward and downward
directions as shown in FIG. 5, and emits the light flux 22 having a
vertically linearly symmetric shape with respect to the
illuminative light axis 18. FIG. 6 is a diagram concretely showing
the light flux 22 refracted by the wedge prism 13 in two vertical
directions. In this case, the light flux 22 refracted in two
vertical directions by the wedge prism 13 is kept to be
parallel.
[0081] The condenser 14 condenses the light flux 22 having two
directions from the wedge prism 13 (parallel light beam inclined at
a predetermined angle with respect to the illuminative light axis
18) on different places (two places of upper and lower places) on
the slit 15 plane to project the image of the light source 11.
[0082] The slit 15 has three types of openings 23, 24, 25 having
different shapes as described later in detail and as shown, for
example, In FIGS. 7A, 8A, 9A in the same manner as in the first
embodiment.
[0083] Since the configuration other than the above-described
configuration is similar to that of the first embodiment, detailed
description is omitted.
[0084] In this case, when the slit 15 is moved, the position of the
slit image can be adjusted on the emission pupil surface of the
objective lens 4. That is, when the slit image on the emission
pupil surface is moved into a fluorescent illumination region 28
shown in FIG. 7B, usual fluorescent observation is possible. When
the slit image is moved into the total reflection region 27, the
total reflection fluorescent observation is possible. Furthermore,
when the slit image is moved in the total reflection region 27, an
incidence angle of the illuminative light upon the specimen from
the objective lens can be finely adjusted, and an oozing depth of
the evanescent light may also be controlled in accordance with an
observation position of the specimen.
[0085] The similar effect and advantage can be obtained to the
first embodiment even when the opening is formed at only either of
upper or lower portion.
[0086] The wedge prism 13 refracts the parallel light beam from the
collector lens 12 in two vertical directions as described above to
emit the light flux 22 (light source image) having a vertically
linearly symmetric shape with respect to the illuminative light
axis 18. The light flux 22 is condensed onto two places of upper
and lower places on the slit 15.
[0087] In this case, as the slit 15, it is preferable to use the
slit in which crescent openings 23 are formed in two places of
upper and lower places having point symmetry with respect to a
center of the slit 15 on the slit 15 plane as shown in FIG. 7A.
[0088] Moreover, the light source images 11a are projected as the
image of the light source 11 in the crescent openings 23 of the
slit 15 as shown in FIG. 7C.
[0089] FIG. 7B shows the image of the slit 15 projected on the
emission pupil surface of the objective lens 4, that is, a slit
image 23a. In FIG. 7B, an orbicular portion shown by meshes in a
pupil diameter 4a of the objective lens 4 shows the total
reflection region 27 where the light is totally reflected by the
boundary surface between the specimen 3 and the cover glass 7. A
shown white circular portion inside the orbicular portion shows the
fluorescent illumination region 28 where the total reflection is
not performed.
[0090] Accordingly, all the light beams transmitted through the
openings 23 of the slit 15 fall in the total reflection region 27
to form the totally reflected light beams. Since the subsequent
operation is similar to that of the first embodiment, detailed
description is omitted.
[0091] A relation between the slit image 23a by the crescent
openings 23 of the slit 15 and the emission pupil surface of the
objective lens 4 will be described in further detail with reference
to FIG. 7B.
[0092] FIG. 7B shows a state in which the center of a slit image
23a by the crescent opening 23 deviates from that of a pupil
diameter 4a of the objective lens 4. As causes for the deviation, a
shift of an optical axis of the optical illumination system to the
FS projection lens 17 from the light source 11, inclination error
of a reflection surface in the dichroic mirror 212, mechanical
eccentricity of the objective lens 4 and the like are
considered.
[0093] In consideration of a case where the slit image 23a has the
same size as that of the total reflection region 27, when the slit
image 23a is eccentric even slightly, a part of the slit image 23a
enters the fluorescent illumination region 28, light leak occurs,
and accordingly a drop of contrast is sometimes caused. However,
when the size of the slit image 23a is set to be smaller than that
of the total reflection region 27, the slit image 23a constantly
stays in the total reflection region 27 even with slight movement
of the slit image 23a for the above-described causes. Therefore,
there is not fear that the light beam enters the fluorescent
illumination region 28 and leaks, and the observation with good
contrast is possible. Therefore, the crescent opening 23 shown in
FIG. 7A is formed in a smaller shape so as not to protrude on a
fluorescent illumination region 28 side even when the slit image
23a slightly moves by magnification errors of the optical
illumination system to the FS projection lens 17 from the light
source 11 and the magnification error of the objective lens 4.
[0094] Moreover, when the cassette 19 is rotated to insert or
remove the mirror units 18a, 18b having different wavelength
characteristics on the observation optical axis 6, a positioning
reproduction accuracy of a rotation direction of the cassette 19
sometimes results in the error of the inclination of the dichroic
mirror 212 or the inclination error of the dichroic mirror 212 for
each of the mirror units 18a, 18b. Moreover, these errors appear as
positional shifts of projection of the slit image 23a in the
objective lens emission pupil surface. In this case, when the slit
image 23a is configured to change its direction in a left-to-right
direction with respect to these errors in FIG. 7B, the slit image
23a by the crescent opening 23 long in the horizontal direction
constantly stays in the total reflection region 27. Accordingly,
the influence of deterioration of contrast can be eliminated with
respect to vibration of the slit image 23a in the horizontal
direction. Even when the position of the opening 23 of the slit 15
slightly shifts with respect to the light source image 11a shown in
FIG. 7C, much light can be taken in from the light source 11,
because the crescent openings 23 has a crescent shape long in the
horizontal direction.
[0095] It is to be noted that the slit 15 in which the crescent
openings 23 are formed has been described above in detail. However,
the present invention is not limited to this. As described in the
first embodiment, for example, a slit in which small-diameter
openings 24 are formed in two positions of upper and lower
positions having point symmetry with respect to the center of the
slit 15 on the slit 15 plane as shown in FIG. 8A, a slit in which
annular openings 25 are formed along the peripheral edge portion of
the slit 15 plane as shown in FIG. 9A, a slit in which elliptic
opening (not shown) are formed and the like are considered.
[0096] In the slit 15 having the small-diameter openings 24, as
shown in FIG. 8C, the light source images 11a by the light source
11 are projected with respect to the openings 24. As shown in FIG.
8B, slit images 24a by the small-diameter openings 24 are projected
on the emission pupil surface of the objective lens 4. In the slit
15 having the small-diameter openings 24, the light source images
11a sometimes shift from the small-diameter openings 24 and are
darkened by deviation of the positions of the openings 24 and the
light source images 11a by the above-described factors. However, if
the light source images 11a greatly decentered, because the opening
24 is hardly extended to the fluorescent illumination area, the
contrast can be prevented being degraded. Since the shape of the
slit 15 is simple, the slit is characterized in that the slit is
easily worked and is inexpensive.
[0097] In the slit 15 having the annular openings 25, as shown in
FIG. 9C, the light source image 11a by the light source 11 is
projected with respect to the openings 25. As shown in FIG. 9B, a
slit image 25a by the openings 25 is projected on the emission
pupil surface of the objective lens 4. When the slit image 25a
shifts to the right/left in the slit 15 having the annular openings
25, an inner diameter of the slit image 25a easily overlaps with
the fluorescent illumination region 28, and the contrast easily
drops. However, even when the annular openings 25 and the light
source image 11a slightly shift, a ratio at which the light source
image 11a deviates from the openings 25 is small. Therefore, this
is effective means for securing the brightness in a case where the
eccentricity of the optical illumination system is small.
[0098] Since the shape of the opening can be matched with that of
the luminescent spot of the general arc light source in the slit 15
having the elliptic openings shown in FIG. 8D and FIG. 8E, it is
possible to efficiently take in the illuminative light.
[0099] Next, the present embodiment is similar to the first
embodiment in a case where the total reflection fluorescent
observation is changed to the usual fluorescent observation to
perform the observation, and therefore the description is
omitted.
[0100] As described above, according to the second embodiment, an
effect similar to that of the first embodiment can be obtained.
[0101] It is to be noted that the size or shape of the light source
11 is not described in the first and second embodiments, but the
size or shape of the light source 11 can be set as follows.
[0102] When the slit 15 has the crescent openings 23 shown in FIG.
7A or the annular openings 25 shown in FIG. 9A, a light source for
obtaining an elliptic light source image 11b as shown in FIGS. 7D
and 9D is used as the light source 11. Moreover, the elliptic light
source image 11b is projected on each slit 15 in a state in which
the longitudinal direction is positioned transversely as shown in
FIGS. 7D and 9D. Then, since the light source image 11b is
projected on a broad range of the opening 23 (25), an illumination
efficiency can further be improved. To embody this, a whole lamp
house 30 in which the light source 11 is stored may be configured
so as to be rotatable around the illuminative light axis 18 in
accordance with the lamp shape of the light source 11. At this
time, the whole lamp house 30 may be rotatably supported, rotated
by a predetermined angle in this state, and fixed via screws.
Needless to say, instead of rotating the lamp house 30, the light
source 11 itself may be rotated in the lamp house 30.
MODIFICATION OF SECOND EMBODIMENT
[0103] Next, a modification of the second embodiment will be
described.
[0104] The modification of the second embodiment is an example in
which the illumination efficiency is raised without using the wedge
prism, and will be described with reference to FIGS. 10A and
10B.
[0105] As shown in FIG. 10A, the light source 11 is movable
vertically in an arrow direction along the plane crossing the
illuminative light axis 18 at right angles. Moreover, the light
source 11 can be positioned in two positions including a position
on the illuminative light axis 18 and a lower position deviating
slightly from the illuminative light axis 18.
[0106] To perform the total reflection fluorescent observation, the
light source 11 is set in a position denoted with reference numeral
11' slightly deviating from the illuminative light axis 18. Then,
as shown in FIG. 10B, the light beams from the light source 11' are
formed into the parallel light beam having a predetermined angle
with respect to the illuminative light axis 18 by the collector
lens 12, and are projected as a light source image 11a' in the
opening 23 in the upper part of the slit 15. Accordingly, the
illumination efficiency can be raised without using the wedge
prism. To return to the usual fluorescent illumination, the light
source 11 may be positioned on the illuminative light axis 18.
[0107] In the present modification, the light source 11 may be
moved in the vertical direction of the light source 11 with one
touch. However, the light source 11 usually has a centering
function. Therefore, when the centering function is used, the
illumination efficiency can be enhanced simply and inexpensively.
It is to be noted that with the use of the wedge prism, less light
is rejected by the collector lens 12 and condenser 14. Therefore,
the illumination efficiency is better that that of the present
modification, but brightness is to be enhanced inexpensively. In
this case, the present modification is effective means.
THIRD EMBODIMENT
[0108] Next, a third embodiment of the present invention will be
described.
[0109] In the third embodiment, means for further reducing the
illumination unevenness is added to the configuration of the second
embodiment.
[0110] FIG. 11 is a diagram showing a schematic configuration of
the third embodiment.
[0111] In the third embodiment, a conical prism 31 is used instead
of the wedge prism 13 described in the second embodiment. In the
conical prism 31, a conical concave portion 31a is formed in the
surface on a light source 11 side, and a surface on a specimen 3
side is formed in a flat surface 31b. Moreover, the conical prism
31 is disposed in such a manner that a vertex of the illuminative
light axis 18 matches that of the conical concave portion 31a on
the optical path of the parallel light beam from the collector lens
12.
[0112] The conical prism 31 refracts the parallel light beam from
the collector lens 12 while keeping a parallel light flux toward
the outside from the illuminative light axis 18 to emit a light
flux 32. FIG. 12 is a diagram concretely showing the light flux 32
refracted toward the outside from the illuminative light axis 18 by
the conical prism 31. Unlike the wedge prism 13, an inner diameter
of the light flux 32 is conical.
[0113] The condenser 14 and slit 15 are disposed in the optical
path of the light flux 32 reflected by the conical prism 31. As the
slit 15, a slit is used in which the annular opening 25 is formed
along the peripheral edge portion as shown in FIG. 13.
[0114] In the configuration, the parallel light beam emitted from
the collector lens 12 is refracted toward the outside from the
illuminative light axis 18 by the conical prism 31. The refracted
parallel light beam is condensed in the annular opening 25 of the
slit 15 by the condenser 14, and projected as the light source
image 11a in the opening 25 of the slit 15. In this case, an
infinite number of the light source images 11a projected on the
annular opening 25 of the slit 15 are projected along a
circumferential direction of the opening 25 around the illuminative
light axis 18 as shown in FIG. 13.
[0115] Moreover, the light transmitted through the slit 15 is
projected as a slit image on the emission pupil surface of the
objective lens 4 via the FS projection lens 17. Accordingly, the
total reflection fluorescent observation is possible in the same
manner as in the second embodiment.
[0116] Thereafter, the light flux 32 refracted toward the outside
from the illuminative light axis 18 is generated by the conical
prism 31, and accordingly the light source image 11a can be
projected along the annular opening 25 of the slit 15. Accordingly,
in addition to the effect of the second embodiment, since the
annular opening 25 can be uniformly illuminated, the illumination
unevenness can be largely reduced.
MODIFICATION OF THIRD EMBODIMENT
[0117] Next, a modification of the third embodiment will be
described
[0118] The modification of the third embodiment is an example
including another means for reducing the illumination unevenness,
and will be described with reference to FIGS. 14A and 14B.
[0119] In this case, the wedge prism 13 is disposed in the optical
path between the collector lens 12 and the condenser 14 in the same
manner as in the second embodiment. Moreover, a slit in which the
annular opening 25 is formed along the peripheral edge portion as
shown in FIG. 14B is used as the slit 15. Further-more, in this
state, the wedge prism 13 is rotated at a high speed in a direction
of an arrow 33 using the illuminative light axis 18 which is a
rotational center. In this case, the prism is rotated once at about
30 msec in the visual observation, or rotated at a rotation number
higher than a scanning speed of photo-detection, when
photo-detection means such as CCD. Accordingly, as shown in FIG.
14B, the light source image 11a rotates along the annular opening
25 around the illuminative light axis 18. Therefore, when a time
average of the rotation is taken, an effect similar to that with
the use of the conical prism 31 described in the third embodiment
is obtained. In this case, the rotation means of the wedge prism 13
can be realized using a known motor and bearing.
[0120] Moreover, the light transmitted through the slit 15 is
projected on the slit image on the emission pupil surface of the
objective lens 4 via the FS projection lens 17. Accordingly, the
total reflection fluorescent observation is possible in the same
manner as in the second embodiment.
[0121] Therefore, when the wedge prism 13 is configured so as to be
rotatable around the illuminative light axis 18 of the optical
illumination system at a high speed, the light source image 11a can
be projected along the annular opening 25, and therefore the total
reflection fluorescent observation is realized with the
illumination without any directionality or unevenness. The cost can
also be reduced without using the expensive conical prism 31.
FOURTH EMBODIMENT
[0122] A fourth embodiment of the present invention will be
described.
[0123] FIG. 15 is a diagram showing a schematic configuration of
the fourth embodiment, and the same components as those of FIG. 1
are denoted with the same reference numerals.
[0124] In the fourth embodiment, six LEDs 34 having micro
luminescent spots are disposed instead of the light source 11. In
this case, six LEDs 34 are disposed in the position of the point
symmetry with respect to the illuminative light axis 18 on the
plane crossing the illuminative light axis 18 at right angles. In
the fourth embodiment, the wedge prism 13 is not required. As the
slit 15, a slit is used in which the crescent openings 23 are
formed in two positions of upper and lower positions of the point
symmetry with respect to the center of the slit 15 on the slit 15
plane as shown in FIG. 16.
[0125] In the above-described configuration, the illuminative
lights emitted from six LEDs 34 are formed into parallel light
beams having a predetermined angle with respect to the illuminative
light axis 18 by the collector lens 12, and are projected as LED
images 35 in the upper/lower crescent openings 23 of the slit 15 by
the condenser 14 as shown in FIG. 16.
[0126] Moreover, the light transmitted through the slit 15 is
projected as the slit image on the emission pupil surface of the
objective lens 4 via the FS projection lens 17. Accordingly, the
total reflection fluorescent observation is possible in the same
manner as in the second embodiment.
[0127] Therefore, since the respective LED images 35 of six LEDs 34
can be projected on accordance with the upper/lower crescent
openings 23 of the slit 15, the illumination efficiency can be
further enhanced.
[0128] It is to be noted that when the slit having the annular
openings 25 as shown in FIG. 17 is used as the slit 15, a large
number of LEDs 34 are arranged in an annular form around the
illuminative light axis 18. Moreover, the lights from the LEDs 34
arranged in the annular form are projected as the LED images 35 in
the annular openings 25 of the slit 15 via the collector lens 12
and condenser 14.
[0129] Since the LED images 35 from the annularly arranged LEDs 34
can be uniformly projected along the annular openings 25 of the
slit 15 in this manner, the illumination unevenness can be
reduced.
[0130] It is to be noted that when the number of LEDs 34 is further
increased and a large number of LEDs are arranged around the
illuminative light axis 18, the LEDs 34 can only be selectively lit
in accordance with the shapes of the openings of the slit 15 to
project the LED images 35 in accordance with various openings. All
the LEDs 34 may be lit in the usual fluorescent illumination
observation.
[0131] According to the fourth embodiment, by the use of the light
source having a plurality of micro luminescent spots arranged to
fill the openings 23 (25) of the slit 15, the light source image is
projected only in a range passing through the openings 23 (25), and
the illuminative light is not introduced except the total
reflection region. Therefore, the illuminative light can be
efficiently taken in, and the total reflection fluorescent
observation is possible with the sufficient brightness and good
contrast. Since the expensive wedge prism or conical prism is not
used, the microscope is inexpensive. Especially, when a slit having
the annular opening 25 is used as the slit 15, the total reflection
fluorescent illumination having remarkably little unevenness may
also be obtained.
FIFTH EMBODIMENT
[0132] A fifth embodiment of the present invention will be
described.
[0133] FIG. 18 is a diagram showing a schematic configuration of
the fifth embodiment, and the same components as those of FIG. 1
are denoted with the same reference numerals.
[0134] In the fifth embodiment, an afocal converter 36 is disposed
as magnification varying means for raising light source
magnification in the optical path between the collector lens 12 and
the wedge prism 13. The afocal converter 36 comprises a convex lens
36a and concave lens 36b. By the afocal converter 36, the parallel
light beam from the collector lens 12 is condensed onto the convex
lens 36a, and diverted by the concave lens 36b. Accordingly, the
parallel light beam whose light source magnification has been
raised can be emitted. The afocal converter 36 is detachably
inserted together with the wedge prism 13 and slit 15 with respect
to the optical path.
[0135] In the slit 15, a slit in which the crescent openings 23 are
formed in the two positions of upper and lower positions of the
point symmetry with respect to the center of the slit 15 on the
slit 15 plane as shown in FIGS. 19A and 19B, or a slit in which the
annular openings 25 are formed along the peripheral edge portion of
the slit 15 as shown in FIGS. 20 and 20B are used.
[0136] When the total reflection fluorescent illumination is
performed, the illuminative light emitted from the light source 11
is projected as a light source image 37 on the slit 15 plane via
the collector lens 12, the convex lens 36a and concave lens 36b
configuring the afocal converter 36, the wedge prism 13, and the
condenser 14. In this case, in the light source image 37, since the
light source magnification is raised by the afocal converter 36,
the light source image 37 projected on the slit 15 plane spreads
sufficiently in a broad range on the respective openings 23, 25 as
shown in FIG. 19B or 20B. FIG. 19A or 20A shows a case where the
afocal converter 36 is not disposed, and the light source image 37
projected on the slit 15 plane overlaps with a part of the openings
23, 25.
[0137] Therefore, to perform the total reflection fluorescent
illumination, when the convex lens 36a and concave lens 36b
configuring the afocal converter 36 are inserted in the optical
path to raise the magnification of the light source image 37, more
openings 23 (25) of the slit 15 can be filled with the light source
images 37, and therefore the illumination efficiency can be further
enhanced.
[0138] The light transmitted through the slit 15 is projected as
the slit image on the emission pupil surface of the objective lens
4 via the FS projection lens 17, and the total reflection
fluorescent observation is possible in the same manner as in the
second embodiment.
[0139] When the small-diameter openings 24 are formed in two
positions of upper and lower positions of the point symmetry with
respect to the center of the slit 15 in the slit 15 as shown in
FIG. 21, even the light source images 37 projected onto the slit 15
plane sufficiently fill the openings 24 of the slit 15 in a state
free of the afocal converter 36. Therefore, even when the light
source magnification is raised particularly using the afocal
converter 36, an effect of enhancement of the illumination
efficiency is little.
[0140] Next, a case where usual fluorescent observation is
performed will be described.
[0141] As shown in FIG. 22, the afocal converter 36, wedge prism
13, and slit 15 are removed from the optical path of the optical
illumination system, and the aperture stop (AS) 29 is inserted
instead of the slit 15. When the wedge prism 13 or the slit 15
enters the optical path, the illumination efficiency drops or the
illumination unevenness increases in the same manner as in the
second embodiment. Therefore, to perform the usual fluorescent
observation, the prism or the slit is removed from the optical
path, and the aperture stop (AS) 29 for adjusting the brightness is
inserted instead of the slit 15. This can be realized by the use of
an inserting/detaching mechanism such as a known slider. The convex
lens 36a and concave lens 36b configuring the afocal converter 36
are effective for enhancing the illumination efficiency. However,
on the contrary, the illumination field is narrowed, and an
observable range is narrowed. Therefore, the afocal converter 36 is
not required at the time of the usual fluorescent observation with
the sufficient brightness, and is also removed from the optical
path.
[0142] It is to be noted that the afocal converter 36 is used to
changing the light source magnification. Therefore, even when the
converter is removed from the optical path, a projection plane of
the light source 11 is unchanged. Therefore, even when the aperture
stop (AS) 29 is inserted in the optical path instead of the slit
15, the light source image is projected on the aperture stop (AS)
29 plane. Therefore, there is no fear that the illumination
unevenness occurs also at the usual fluorescent observation time,
the illumination efficiency does not drop, the illumination is
bright, and therefore optimum microscopic inspection can be
performed.
[0143] In this state, specimen can be visually observed by the
generally known usual fluorescent observation method.
[0144] It is to be noted that another variable magnification lens
is also usable as means for raising the light source magnification
in addition to the afocal converter 36.
[0145] In the fifth embodiment, the afocal converter 36 is disposed
as magnification varying means for raising a projection
magnification of the light source 11 between the slit 15 and the
light source 11. Accordingly, in the total reflection illumination,
the magnification of the light source image is raised, and even a
portion incapable of filling the openings 23 of the slit 15 is
filled with the light source image 37. Accordingly, the total
reflection fluorescent observation is possible by brighter
illumination. Even when the afocal converter 36 is inserted or
removed with respect to the optical path, the projection position
of the light source image 37 in the optical-axis direction does not
change. Therefore, the illumination efficiency at a total
reflection fluorescent observation time does not drop. Moreover,
the illumination unevenness does not easily occur at a usual
fluorescent observation time, and optimum microscopic inspection
can be performed in each observation.
SIXTH EMBODIMENT
[0146] A sixth embodiment of the present invention will be
described.
[0147] FIG. 23 is a diagram showing a schematic configuration of a
main part of a sixth embodiment, and the same components as those
of FIG. 1 are denoted with the same reference numerals.
[0148] In this case, the collector lens 12, condenser 14, and slit
15 are disposed on the illuminative light axis 18 of the light from
the light source 11. A convex lens 44 having a large diameter is
disposed as a lens having a weak refractive power between the
collector lens 12 and the condenser 14.
[0149] The convex lens 44 is disposed while a central axis 44a is
largely shifted from the illuminative light axis 18, and the
parallel light beam from the collector lens 12 is refracted by a
predetermined angle with respect to the illuminative light axis 18.
The condenser 14 condenses a light flux 45 refracted by the convex
lens 44 is condensed on the slit 15 plane, and the light source
image 11a is projected. Also in this case, a slit is used in which
a crescent opening 43 is formed on the slit plane is used in the
same manner as in FIG. 3A. The light source image 11a is projected
on the crescent opening 43a via the condenser 14.
[0150] Also in the sixth embodiment, the convex lens 44 and slit 15
can be detachably inserted with respect to the optical path of the
illuminative light by known switching mechanisms such as a slider.
The slit 15 inserted in the optical path is movable further along
the plane crossing the illuminative light axis 18 at right angles
in a known arrow direction.
[0151] The other configuration is similar to that of FIG. 1.
[0152] Therefore, the effect similar to that of the first
embodiment can be expected.
SEVENTH EMBODIMENT
[0153] A seventh embodiment of the present invention will be
described.
[0154] FIG. 24 is a diagram showing a schematic configuration of a
main part of the seventh embodiment, and the same components as
those of FIG. 1 are denoted with the same reference numerals.
[0155] In FIG. 24, the collector lens 12, condenser 14, and slit 15
are disposed on the illuminative light axis 18 of the light from
the light source 11. A parallel plane plate 46 is disposed between
the condenser 14 and the slit 15.
[0156] The parallel plane plate 46 is inclined at a predetermined
angle with respect to the illuminative light axis 18 and disposed
to move the light beam from the condenser 14 in parallel with the
illuminative light axis 18, and condenses the light on the slit 15
plane to project the light source image 11a. Also in this case, a
slit in which the crescent opening 43 is formed on the slit plane
is used as the slit 15 in the same manner as in FIG. 22. The light
source image 11a is projected on the crescent opening 43 via the
condenser 14.
[0157] Also in this case, the parallel plane plate 46 and slit 15
are detachably inserted with respect to the optical path of the
illuminative light by known switching mechanisms such as the
slider. Moreover, the inclination angle of the parallel plane plate
46 inserted in the optical path is adjustable in the arrow
direction. Therefore, even when the total reflection region differs
with the type of the objective lens, the inclination angle of the
parallel plane plate 46 can be adjusted to adjust the light source
image 11a in an optimum position in the total reflection region.
Furthermore, even the slit 15 inserted in the optical path can move
along the plane crossing the illuminative light axis 18 at right
angles in the arrow direction.
[0158] The other configuration is similar to that of FIG. 1.
[0159] Therefore, the effect similar to that of the first
embodiment can be expected.
EIGHTH EMBODIMENT
[0160] An eighth embodiment of the present invention will be
described.
[0161] FIG. 25 is a diagram showing a schematic configuration of a
main part of the eighth embodiment, and the same components as
those of FIG. 1 are denoted with the same reference numerals.
[0162] In FIG. 25, the collector lens 12, condenser 14, and slit 15
are disposed on the illuminative light axis 18 of the light from
the light source 11. Mirrors 47, 48 are disposed between the
condenser 14 and the slit 15.
[0163] The light beam from the condenser 14 is reflected by the
mirror 47, and the reflected light is reflected by the mirror 48.
Accordingly, the optical path of the optical illumination system is
moved in parallel, that is, the light beam from the condenser 14 is
moved in parallel with the illuminative light axis 18 and condensed
onto the slit 15 plane to project the light source image 11a. Even
in the eighth embodiment, the slit in which the crescent opening 43
is formed on the slit plane as described with reference to FIG. 22
is used as the slit 15. The light source image 11a is projected on
the crescent opening 43 via the condenser 14.
[0164] Even in this case, the mirrors 47, 48 and the slit 15 can be
detachably inserted with respect to the optical path of the
illuminative light by the known switching mechanisms such as the
slider. In the mirrors 47, 48, the mirror 48 is movable in the
arrow direction, and the distance from the mirror 47 can be
adjusted. Accordingly, even when the total reflection region
differs with the type of the objective lens, the distance between
the mirrors 47, 48 can be adjusted so as to adjust the light source
image 11a in an optimum position in the total reflection region.
Furthermore, even the slit 15 inserted in the optical path can move
along the plane crossing the illuminative light axis 18 at right
angles in the arrow direction.
[0165] The other configuration is similar to that of FIG. 1.
[0166] Therefore, the effect similar to that of the first
embodiment can be expected.
[0167] The present invention is not limited to the above-described
embodiments, and can be variously modified in a range in which the
scope is not changed. For example, in the embodiments, the optical
device for decentering the optical axis has been described.
Alternatively, a plurality of optical devices having different
eccentricities of the optical axis are prepared, and may also be
selectively used in accordance with the type of the objective lens
(the total reflection region differs). The plane of the slit 15 on
the light source 11 side may also be formed in a reflection plane
or an irregular reflection plane. In this case, degradation by heat
or the like in a portion which interrupts the light beam of the
slit 15, that is, a portion irradiated with the light beam can be
reduced. Furthermore, in the embodiments, the inverted microscope
has been described by which the observation is performed by the
objective lens disposed under the specimen, but a transmission
illumination type may also be used in which the total reflection
fluorescent illumination is performed using a condenser lens, or an
erected microscope may also be used.
[0168] Furthermore, the embodiments include various stages of
inventions, and various inventions can be extracted by an
appropriate combination of a plurality of configuring elements. For
example, even when several configuring elements are removed from
all the configuring elements described in the embodiments, the
described problems to be solved by the present invention can be
resolved, and the described effects of the present invention are
obtained. In this case, the configuration from which the
configuring elements are removed can be extracted as the
invention.
[0169] It is to be noted that the above-described embodiments also
include the following inventions.
[0170] A fluorescence microscope according to a first aspect of the
present invention is characterized by comprising a light source; an
optical illumination system which forms an optical path to
irradiate a specimen with a light beam from the light source; an
objective lens which condenses the light beam of the optical
illumination system onto the specimen; an optical device which is
disposed on the optical path of the optical illumination system and
which decenters the light beam by decentering an optical axis of
the optical path; and a slit which passes the light beam decentered
by the optical device through a total reflection illumination
region on an emission pupil surface of the objective lens. In the
first aspect, the following manners are preferable.
[0171] (1) The total reflection illumination is illumination using
an evanescent light oozing by a predetermined amount on a specimen
side in a boundary surface between glass on which the specimen is
laid and the specimen.
[0172] (2) The optical device and the slit are movable along a
plane vertical to the optical axis of the optical path of the
optical illumination system.
[0173] (3) The opening has at least one of a crescent shape, a
circular shape, a half-ring shape, and an elliptic shape.
[0174] (4) The optical device is a prism.
[0175] (5) In (4), the prism is either a wedge prism or a conical
prism.
[0176] (6) In (4), the prism is a wedge-shaped plane plate.
[0177] (7) The optical device is a lens having a small refractive
index.
[0178] (8) The optical device is a parallel plane plate disposed
with a predetermined angle with respect to the optical axis.
[0179] (9) The optical device comprises a pair of mirrors which
move the optical path of the optical illumination system.
[0180] (10) The optical device includes a plurality of optical
devices, and the plurality of optical devices are selectively
inserted in the optical path of the optical illumination
system.
[0181] (11) In (5) or (6), the wedge prism is rotatable around the
optical axis of the optical illumination system.
[0182] (12) The light source has a luminescent spot in which a
projected image on an emission pupil surface of the objective lens
is smaller than an emission pupil diameter of the objective
lens.
[0183] (13) In (12), the light source is either a micro arc type
light source or a light source comprising a plurality of micro
luminescent spots.
[0184] (14) In (13), the light source comprising the plurality of
micro luminescent spots includes a plurality of light emitting
diodes.
[0185] (15) In (14), the plurality of light emitting diodes are
arranged on a circumference having a predetermined diameter.
[0186] (16) An optical magnification varying system which is
disposed in the optical path of the optical device on the light
source side to raise a projection magnification of the light source
is further provided.
[0187] (17) In (16), the optical magnification varying system
includes an afocal converter.
[0188] (18) The slit has a reflection surface or an irregular
reflection surface formed on a plane on a light source side.
[0189] A fluorescence microscope according to a second aspect of
the present invention is characterized by comprising: a light
source; an optical illumination system which forms an optical path
to irradiate a specimen with a light beam from the light source; an
objective lens which condenses the light beam of the optical
illumination system onto the specimen; and a slit which passes the
light beam from the light source through a total reflection
illumination region on an emission pupil surface of the objective
lens, in which an emission position of the light beam emitted from
the light source is movable between an optical axis of the optical
illumination system and a position shifting from the optical axis
by a predetermined distance.
[0190] According to the embodiments of the present invention, the
optical device for decentering the optical axis of the optical path
is disposed between the light source and the slit. Accordingly, the
illuminative light can be efficiently taken in the total reflection
region having an emission pupil diameter of the objective lens by
the eccentricity of the optical axis of the optical path.
[0191] Moreover, the optical device and slit are detachably
inserted in the optical path. Therefore, when the states are only
selected, the illuminative light can be efficiently taken into the
total reflection region of the emission pupil surface of the
objective lens or the fluorescent illumination region where the
total reflection is not performed.
[0192] Furthermore, the slit for performing the total reflection
illumination has the crescent opening which passes the light beam
only through a part of the total reflection region. Accordingly,
even when the position or the size of the slit image changes by the
eccentricity or the magnification error of the optical illumination
system, the slit image can be prevented from deviating from the
total reflection region of the emission pupil surface of the
objective lens.
[0193] Moreover, since the wedge prism or the conical prism is used
as the optical device, the respective parallel light beams in the
light flux can be refracted by an equal angle, little aberration is
generated by the prism or the condenser, and a satisfactory light
source image can be projected on the slit plane.
[0194] Furthermore, since a light source having a plurality of
micro luminescent spots is used as the light source, the light
source image is projected only in a range passing through the slit.
Since the illuminative light is not introduced into a region other
than the total reflection region, the illuminative light can be
efficiently taken.
[0195] Moreover, since the wedge prism is rotatable around the
optical axis of the optical illumination system, the light source
image can be projected on the annular shape, and the illumination
is obtained without any directionality or unevenness.
[0196] Furthermore, since an optical magnification varying system
for changing a projection magnification is disposed in the optical
path of the optical device on the light source side, in the total
reflection illumination, the magnification of the light source
image is raised, and even the portion of the slit that cannot be
filled is filled with the light source image, and the total
reflection fluorescent observation by brighter illumination is
possible.
[0197] As described above, according to the embodiments of the
present invention, there can be provided a total reflection
fluorescent microscope in which use efficiency of the illuminative
light is raised, and the total reflection fluorescent observation
is possible with the sufficient brightness and good contrast.
[0198] Additional advantages and modifications will readily occur
to those skilled in the art. Therefore, the invention in its
broader aspects is not limited to the specific details and
representative embodiments shown and described herein. Accordingly,
various modifications may be made without departing from the spirit
or scope of the general invention concept as defined by the
appended claims and their equivalents.
* * * * *