U.S. patent application number 11/882153 was filed with the patent office on 2007-12-27 for endoscopic, in vivo cellular observation methods.
Invention is credited to Naoki Hasegawa.
Application Number | 20070299312 11/882153 |
Document ID | / |
Family ID | 33156631 |
Filed Date | 2007-12-27 |
United States Patent
Application |
20070299312 |
Kind Code |
A1 |
Hasegawa; Naoki |
December 27, 2007 |
Endoscopic, in vivo cellular observation methods
Abstract
An endoscope observation system for in vivo cellular observation
is disclosed that includes an illumination optical system having a
light source for supplying illumination light to an object, an
objective optical system that forms a magnified image of the object
such that the absolute value of the image scale factor exceeds
unity, and an image pickup unit that detects the magnified image.
The illumination optical system is provided with a wavelength
selection means for dividing, among the blue, green, and red
wavelength ranges in the illumination light from the light source,
either the blue wavelength range or the red wavelength range into
two wavelength bands T1 and T2, with the wavelength band T1 being
nearer the green wavelength range than is the wavelength band T2,
and light in the wavelength band T1 is prevented from illuminating
the object. An in vivo cellular observation method is also
disclosed using an endoscope.
Inventors: |
Hasegawa; Naoki; (Chofu,
JP) |
Correspondence
Address: |
ARNOLD INTERNATIONAL
P. O. BOX 129
GREAT FALLS
VA
22066-0129
US
|
Family ID: |
33156631 |
Appl. No.: |
11/882153 |
Filed: |
July 31, 2007 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10808309 |
Mar 25, 2004 |
7267648 |
|
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11882153 |
Jul 31, 2007 |
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Current U.S.
Class: |
600/160 |
Current CPC
Class: |
A61B 1/00188 20130101;
A61B 5/0059 20130101; A61B 5/0084 20130101; A61B 1/05 20130101;
A61B 1/043 20130101; A61B 1/0669 20130101; A61B 1/0646 20130101;
A61B 5/0071 20130101 |
Class at
Publication: |
600/160 |
International
Class: |
A61B 1/06 20060101
A61B001/06 |
Foreign Application Data
Date |
Code |
Application Number |
Mar 31, 2003 |
JP |
2003-094028 |
Claims
1-7. (canceled)
8. An endoscopic, in vivo cellular observation method in which,
based on magnified images of living tissue, the number of cell
nuclei captured in the field of view is used to evaluate the cell
size, or the distance between cell nuclei captured in the field of
view is used to evaluate the population density of the nuclei, for
diagnosis of abnormal cells, said method comprising the following
steps: (a) introducing a coloring agent having blue or red
wavelength absorption into cells of living tissue to be imaged so
as to enhance the contrast of cell nuclei using the difference in
the retention rate of the coloring agent between the nuclei and the
other portions of the cells; (b) illuminating the cells with light
having the absorption wavelength; and (c) displaying several tens
to several hundreds of cell nuclei captured in the field of view so
as to facilitate said diagnosis.
9. An endoscopic, in vivo cellular observation method comprising
the following steps: (a) applying a blue or red wavelength band
absorption substance to cells of living tissue to be imaged; (b)
illuminating, when dividing the absorption wavelength band into two
wavelength bands T1 and T2, the cells with the illumination light
of which the wavelength band T1 closer to the green wavelength
range is cut off, thereby enhancing the contrast of cell nuclei
using the difference in light absorption for the wavelength band T2
between the cell nuclei and other portions of the cells; and (c)
displaying several tens to several hundreds of cell nuclei captured
in the field of view for the purpose of evaluating the likelihood
that the in vivo cells are cancerous.
10. An endoscopic, in vivo cellular observation method in which,
based on magnified images of living tissue, the ratio of the area
of the cell nuclei divided by the area within the cell walls in the
field of view is evaluated for diagnosis of abnormal cells, said
endoscopic, in vivo cellular observation method comprising the
following steps, performed in the order indicated: (a) introducing
a coloring agent having blue or red wavelength absorption into
cells of living tissue to be imaged so that the contrast of cell
nuclei will be enhanced due to a difference in retention rate of
the coloring agent in the cell nuclei versus other portions of the
cells; (b) illuminating the cells with light having the absorption
wavelength; (c) displaying several cell nuclei captured in the
field of view for the purpose of evaluating the likelihood that the
in vivo cells are cancerous.
11. (canceled)
12. The endoscopic, in vivo cellular observation method according
to claim 10, wherein: the endoscope that is used has an objective
optical system with a numerical aperture on the object side of 0.3
or larger.
13-16. (canceled)
Description
[0001] This application claims the benefit of foreign priority from
Japanese Patent Application No. 2003-094028, filed Mar. 31, 2003,
the contents of which are hereby incorporated by reference. This is
a divisional application of allowed U.S. application Ser. No.
10/808,309 that was filed Mar. 25, 2004.
BACKGROUND OF THE INVENTION
[0002] Conventional endoscopes have a large field of view that is
in the range of about 90.degree. to 140.degree. so that tissues
inside a body can be observed without overlooking lesions. They
also change the distance to the object in order to obtain either
magnified or reduced-sized images of an object to be observed, and
thus have a large depth of field for a fixed focus point so that
objects at distances between 3 mm and 50 mm can be observed without
refocusing.
[0003] Conventional endoscopes also have an image scale factor with
an absolute value of about 30 to 50 when the image is displayed on
a monitor having a 14-inch screen, which is sufficient to observe
diseased tissues. Zoom optical systems are used in order to obtain
further magnified images, with the absolute value of the image
scale factor being approximately 70 when displayed on a monitor
having a 14-inch screen. The zoom optical system typically has a
built-in, zoom lens driving mechanism. As a result, the endoscope
has an insert tip with an outer diameter that is larger than 10 mm
and requires complex operations. Such endoscopes have limited
applications.
[0004] The manner in which living tissues are observed using a
conventional endoscope will now be described with reference to FIG.
1. Living tissues to be observed by a conventional endoscope often
include a mucous membrane 1, transparent epithelial cells 2 and
underlying parenchymal tissues 3 in which blood vessels run. Light
illumination emitted at the endoscope tip part 4 must first pass
through the mucous membrane and the transparent epithelial cells
before reaching the parenchymal tissues. The illumination light
which reaches the parenchymal tissues 3 is scattered by the
parenchymal tissues 3. Of the light that is scattered by the
parenchymal tissues, most re-enters the epithelial cells. The
illumination light is also scattered by cell walls 5 and cell
nuclei 6 when it is transmitted through the transparent epithelial
cells. The light rays B1, B2 that are scattered by the cell nuclei
of the epithelial cells are weak and thus the light rays that are
scattered by the parenchymal tissues dominate. Consequently, in a
conventional endoscope, only the parenchymal tissues are observed
through an objective optical system.
[0005] When it becomes difficult to provide a diagnosis of an
abnormality by observing images of a tissue, such as when a lesion
is very small, a suspicious-looking tissue may be excised during
the course of an endoscopic examination. The cells of the excised
tissue are then examined under a microscope. Whereas an endoscope
generally uses incident illumination from an illumination optical
system that is positioned around an objective optical system, a
microscope instead generally uses an objective optical system and
an illumination optical system that are positioned on opposite
sides of a sample. The sample is normally pre-processed in order to
make it more suitable for observation, such as by removing the
parenchymal tissues by slicing the sample thin in order to reduce
scattering and/or by staining the sample in order provide better
contrast.
[0006] The manner in which a sample is observed using a microscope
will now be described with reference to FIG. 2. A prepared sample
is fixed onto a cover glass 7 and illuminated from below with light
from an illumination system 8. Illumination light rays A1', A2' are
diffracted by the cell walls and cell nuclei as they transmit
through the sample 9. The diffracted light rays B1', B2' interfere
with one another both constructively and destructively, producing
interference fringes that provide visible contrast. Thus, one can
observe the sample by using an objective optical system 10 placed
above the sample.
[0007] Laser-scanning-type confocal endoscopes which have a
resolution sufficient for cellular observation have been proposed
that may be inserted within a living body. These typically use a
confocal optical system having a pinhole for passing an Airy disk
light pattern at a position that is conjugate to the image plane,
and the confocal optical system thus acquires diffraction-limited
information for each point of an object in the field of view. A
laser beam directed from a light emitting optical system scans the
object, and information obtained from the reflected light from the
object for each point is combined so as to produce an image
representing either a two-dimensional or a three-dimensional
object. High resolution can thus be realized not only within the
image plane, but also in the depth direction.
[0008] It takes from several days to several weeks to identify
abnormal tissue using conventional procedures wherein living
tissues are excised and examined in vitro. Moreover, a cellular
sample that is isolated and fixed for observation is only a tiny
part of a removed tissue. Thus, although a cellular sample provides
information on cellular structures, it is incapable of providing
important functional information, such as information concerning
fluid circulation within cells. This is because the circumstances
between in vitro and in vivo examination are completely different.
Thus, there is a need for magnifying endoscopes that will provide
real-time, in vivo observation of intact living cells.
[0009] In order to form cellular images of a lesion within a living
body, a small-sized image pickup unit is necessary that is provided
with an objective optical system with an image scale factor having
an absolute value that is nearly as high as that of a microscope
and which provides high resolution. The objective optical system
used in a conventional endoscope does not meet these requirements.
As mentioned previously, in a conventional endoscope as shown in
FIG. 1, the illumination light is diffracted by the cell walls and
cell nuclei as it transmits through the epithelial cells. The
diffracted light rays B1, B2 are weak and the light rays A1, A2
that are scattered by the parenchymal tissues are dominant.
Consequently, using a convention endoscope, only data from the
parenchymal tissues is imaged by the objective optical system.
[0010] Although a conventional objective optical system as used in
microscopes is satisfactory as far as providing sufficient imaging
performance, such an objective optical system is too large for easy
insertion into a living body. Laser-scanning-type confocal
endoscopes have a problem in that their scanning speeds are still
too slow for real-time, in vivo observations. Thus, as described
above, an image pickup unit that meets the requirements for in vivo
cellular observation has not yet been realized.
BRIEF SUMMARY OF THE INVENTION
[0011] The present invention relates to a magnifying image pickup
unit suitable for in vivo cellular observation, an endoscope for in
vivo cellular observation, and an in vivo cellular observation
method.
BRIEF DESCRIPTION OF THE DRAWINGS
[0012] The present invention will become more fully understood from
the detailed description given below and the accompanying drawings,
which are given by way of illustration only and thus are not
limitative of the present invention, wherein:
[0013] FIG. 1 is an illustration to explain a principle of living
tissue observation using a prior art endoscope;
[0014] FIG. 2 is an illustration to explain a principle of excised
sample observation using a microscope;
[0015] FIG. 3 shows a configuration of a magnifying image pickup
unit according to the present invention, along with other
components for displaying endoscopic images;
[0016] FIG. 4 shows the spectral wavelength distribution of an
image detected by an image pickup unit according to the present
invention;
[0017] FIG. 5 shows a magnified image of cells that have been
stained for observation according to an embodiment of the present
invention;
[0018] FIG. 6 shows the appearance of cells in the background that
overlap cells of interest in the foreground;
[0019] FIG. 7 shows the spectral transmittance of a wavelength
selection filter used in an embodiment of the present
invention;
[0020] FIG. 8 shows an image in which information from regions
other than regions at a desired depth is eliminated;
[0021] FIG. 9 shows the spectral transmittance of another
wavelength selection filter used in an embodiment of the present
invention;
[0022] FIGS. 10(a) and 10(b) are a side cross section and an end
view, respectively, of an insert tip part of an endoscope imaging
system that uses specific wavelengths for observation;
[0023] FIGS. 11(a) and 11(b) are a cross-sectional view and an end
view, respectively, of the magnifying image pickup unit of
Embodiment 1;
[0024] FIGS. 12(a) and 12(b) are a cross-sectional view and an end
view, respectively, of the magnifying image pickup unit of
Embodiment 2;
[0025] FIG. 13 illustrates an illumination method suitable for in
vivo cellular observation;
[0026] FIG. 14 is a flow chart of the steps for in vivo cellular
observation according to the present invention; and
[0027] FIG. 15 shows an image pickup unit that simultaneously
displays both conventional endoscope images and fluorescent images
for observation of cell nuclei.
DETAILED DESCRIPTION
[0028] A magnifying image pickup unit according to the present
invention that is applied to an endoscope for use in in vivo
cellular observation will now be described.
[0029] First, a conventional endoscope having a wide-angle field of
view is used so as to provide a thorough examination of tissues in
an area of the body without overlooking any area that may contain
diseased tissue. Because it is difficult to diagnose a region of
tissue using images observed with a conventional endoscope, an
endoscope to which the magnifying image pickup unit of the present
invention is applied (hereinafter termed a magnifying endoscope) is
used to cellularly examine a region of tissue.
[0030] For magnified cellular observation, coloring agents
generally will have been previously delivered to the area, if
necessary. Within a certain time period after the coloring agent is
delivered, the difference in the time required for the cell nuclei,
the cell walls, and the other cell components to excrete the
coloring agent creates contrast in the object. Then, a magnifying
endoscope is guided to the region and contact is made with the
object at the tip of the magnifying endoscope while observations
using a conventional endoscope are continued. Preferably, a tissue
image from a conventional endoscope and a cellular image from a
magnifying endoscope are displayed simultaneously on a TV monitor.
In this way, the magnifying endoscope can be guided precisely to a
very small, targeted region within an extensive observation field
of view in order to magnify and observe cell nuclei and cell
walls.
[0031] Before providing a detailed explanation of a magnifying
image pickup unit according to the present invention, the
requirements for an image pickup unit that may be used in a
magnifying endoscope will be discussed. First, the scale factor
required for visualizing fine cellular structures will be
discussed. The overall observation scale factor bm of an image
observed on a display monitor is given by the following equation:
bm=|.beta.o|bd Equation (A) where
[0032] .beta.o is the image scale factor of the objective optical
system, namely, the height of the image as formed on the image
pickup element divided by the actual height of the object, and
[0033] bd is the display scale factor, defined as the monitor
display screen element size divided by the image pickup element
size.
[0034] Conventional endoscopes realize an overall observation scale
factor of about 30 to 50 when the images are viewed on a 14-inch
display monitor. Zoom optical systems having a magnifying function
realize an image scale factor with an absolute value of
approximately 70. However, an overall observation scale factor in
the range of approximately 200 to 2000 is necessary for cellular
observation when viewed on a 14-inch monitor. Therefore, it is
desired that the objective optical system satisfies the following
conditions: 1<|.beta.o|.ltoreq.10 Condition (1) 0.9.ltoreq.|cos
wy'/cos wy|.ltoreq.1.1 Condition (2) where
[0035] .beta.o is the image scale factor of the objective optical
system,
[0036] wy' is the incident angle at which a chief ray corresponding
to the largest field angle enters the image pickup surface, and
[0037] wy is the half-field angle.
[0038] When the lower limit of Condition (2) is not satisfied, the
incident angles of rays onto the image pickup element will be too
large, failing to maintain uniform image qualities (for example,
color reproducibility and brightness) within the field of view.
When the upper limit of Condition (2) is not satisfied, the field
angle will be too large, failing to ensure a required scale
factor.
[0039] Image resolution will now be discussed. Diseased tissues can
be identified with an image resolution in the range of millimeters
and sub-millimeters. However, cellular observations require an
image resolution in the range of microns and sub-microns. In order
to form detailed images of an object that is both transparent and
provides only a small difference in refractive indexes, the
interference of diffracted light rays from the object may be used
so as to provide images having enhanced contrast. The objective
optical system needs to have a larger numerical aperture NA on the
object side so as to collect higher orders of diffracted light rays
and, preferably, satisfies the following Condition (3):
0.1.ltoreq.NA.ltoreq.0.8 Condition (3)
[0040] In the case where the cell walls are observed, it is
preferable to satisfy the following Condition (3'):
0.3.ltoreq.NA.ltoreq.0.8 Condition (3')
[0041] In addition, in order to obtain both high contrast and high
resolution imaging, the objective optical system should have a
resolution higher than that determined by the pitch of the image
pickup element, without exceeding the resolution determined by the
diffraction limit. The objective optical system preferably
satisfies the following Condition (4):
0.1.ltoreq.|pNA.sup.2/(0.61.lamda..beta.o)|.ltoreq.0.8 Condition
(4) where
[0042] p is the pixel size of the image pickup element,
[0043] NA is the numerical aperture of the objective optical system
on the object side;
[0044] .lamda. is the e-line wavelength (i.e., .lamda.=0.546
.mu.m), and
[0045] .beta.o is the image scale factor of the objective optical
system.
[0046] When the lower limit of Condition (4) is not satisfied,
sufficient contrast will not be obtained. When the upper limit of
Condition (4) is not satisfied, aberrations become difficult to
correct, resulting in the failure to obtain fine images.
[0047] Miniaturization (i.e., down-sizing) of the magnifying
endoscope will now be discussed. It is desirable that the
magnifying endoscope have an outer diameter .PHI. of less than 4 mm
in order that it may be guided to an observation object through a
treatment tool insert channel of a conventional endoscope.
Accordingly, it is desirable that the objective optical system be
miniaturized so as to have an outer diameter .PHI. of less than 2
mm.
[0048] A desired, small-sized, objective optical system having an
image scale factor with a large absolute value and a high
resolution comprises, in order from the object side: a lens unit
having positive refractive power and an aperture stop, wherein the
following Condition (5) is satisfied:
0.2.ltoreq..PHI.1/(.PHI.2f1).ltoreq.2 Condition (5) where
[0049] .PHI.1 is the diameter of the aperture stop,
[0050] .PHI.2 is the largest outer diameter of the objective
optical system, and
[0051] f1 is the focal length of the lens unit having a positive
refractive power.
[0052] Condition (5) prevents the objective optical system from
having a larger diameter in association with a larger numerical
aperture NA, thus facilitating miniaturization. When the lower
limit of Condition (5) is not satisfied, the objective optical
system will have a larger total length and a larger maximum
diameter, hampering miniaturization. When the upper limit of
Condition (5) is not satisfied, aberrations become difficult to
correct.
[0053] In order to obtain a flat image surface, it is desirable
that the objective optical system be formed of, in order from the
object side, a front lens unit having positive refractive power, an
aperture stop, and a rear lens unit having positive refractive
power. In such a case, the following Condition (6) is preferably
satisfied in order to achieve both miniaturization and an image
scale factor having a large absolute value:
2.ltoreq.f2/f1.ltoreq.10 Condition (6) where
[0054] f1 is the focal length of the front lens unit, and
[0055] f2 is the focal length of the rear lens unit.
[0056] When the lower limit of Condition (6) is not satisfied, a
required image scale factor having a large absolute value will not
be maintained. When the upper limit of Condition (6) is not
satisfied, a larger overall length and a larger maximum diameter of
the objective optical system will hamper miniaturization.
[0057] Various embodiments of a magnifying image pickup unit of the
present invention will now be described.
EMBODIMENT 1
[0058] The structure of Embodiment 1 of the magnifying image pickup
unit will now be discussed with reference to FIGS. 11(a) and 11(b),
which show a side cross section and an end view, respectively, of
the magnifying image pickup unit according to this embodiment.
[0059] The objective unit comprises an objective lens unit 101
having a uniform diameter in an objective frame 102. The objective
lens unit 101 consists of, in order from the object side: a first
lens group G1 having positive refractive power, an aperture stop
103, and a second lens group G2 having positive refractive power.
An image pickup element 105 is affixed to an image pickup frame 106
via a cover glass 104, thereby forming an image sensor unit.
[0060] The image pickup unit is focused by changing the distance
107 between the objective lens unit and the image sensor unit. An
insert section for a magnifying endoscope is constructed of a hard
tip member 108 and an outer sheath member 110. The image pickup
unit is affixed to the insert section via an intermediate member
109.
[0061] FIG. 11(b) is an end view looking in the direction indicated
by the arrow A in FIG. 11(a). The intermediate member 109 has
cutouts (indicated by cross-hatching) at its periphery through
which an illumination fiber 111 may be inserted and affixed
thereto. After the intermediate member 109 and illumination fiber
are affixed to the hard tip member 108, the image pickup unit is
inserted and affixed to the intermediate member 109.
[0062] Referring to FIG. 11(a), when adjustment is required, for
example, in the absolute value of the image scale factor, the gaps
112a and 112b that are provided before and after the aperture stop
103 can be adjusted to provide a larger or smaller space, if
necessary. To do so, gap adjustment rings that are made of
ultra-thin plates may be used for gap adjustment. A gap adjustment
part is designed to hold a stack of such ultra-thin plates. A
different number of ultra-thin plates may be used according to the
varied gap sizes needed as a result of an assembly process in which
parts having a variety of dimensional errors are used.
[0063] Table 1 below lists the surface number #, in order from the
object side, the radius of curvature R (in mm) of each surface, the
on-axis surface spacing D (in mm), as well as the refractive index
Nd and the Abbe number .upsilon.d (both at the d-line) of each
optical element of Embodiment 1. Also listed is the outer lens
diameter LD of each lens element of Embodiment 1. In the bottom
portion of the Table are listed the distance to the object and the
image height, in mm. TABLE-US-00001 TABLE 1 # R D Nd .upsilon.d LD
1 .infin. 0.46 1.5183 64.14 1 2 0.84 0.17 3 .infin. 0.4 1.7323
54.68 1 4 -0.817 0.05 5 1.353 0.65 1.7323 54.68 1 6 -0.703 0.25
1.7044 30.131 7 -3.804 0.09 8 .infin. (stop) 0.03 9 .infin. 0.4
1.5156 75.00 1 10 .infin. 0.2 11 1.566 0.4 1.67 48.32 1 12 -1.566
0.2 13 -0.729 0.3 1.5198 52.43 1 14 .infin. 0.56 15 .infin. 0.4
1.5183 64.14 16 .infin. 0.01 1.5119 63.00 17 .infin. 0.4 1.6138
50.20 18 .infin. 0.01 1.5220 63.00 19 .infin. 0 Distance to the
object = 0 Image height = 0.500
EMBODIMENT 2
[0064] FIG. 12(a) is a side cross section of the magnifying image
pickup unit of Embodiment 2, and FIG. 12(b) shows an end view
looking in the direction of the arrow A shown in FIG. 12(a).
[0065] Table 2 below lists the surface number #, in order from the
object side, the radius of curvature R (in mm) of each surface, the
on-axis surface spacing D (in mm), as well as the refractive index
Nd, and the Abbe number .upsilon.d (both at the d-line) of each
optical element for Embodiment 2. Also listed is the outer lens
diameter LD of each lens element of Embodiment 2. In the bottom
portion of the Table are listed the distance to the object and the
image height, in mm. TABLE-US-00002 TABLE 2 # R D Nd .upsilon.d LD
1 .infin. 0.88 1.8882 40.76 1.2 2 -0.703 0.05 3 .infin. 0.4 1.5183
64.14 1.2 4 -1.485 0.05 5 2.085 0.76 1.8081 46.57 1.2 6 -0.703 0.25
1.8126 25.42 1.2 7 .infin. 0.05 8 .infin. (stop) 0.03 9 .infin. 0.4
1.5156 75.00 1.2 10 .infin. 0.43 11 1.131 0.5 1.8395 42.72 1.2 12
-3.127 0.2 13 -1.061 0.3 1.8126 25.42 1.2 14 .infin. 0.2 15 -0.592
0.3 1.8081 46.57 1.2 16 2.132 0.77 1.8126 25.42 1.2 17 -1.262 0.77
18 .infin. 0.4 1.5183 64.14 19 .infin. 0.01 1.5119 63.00 20 .infin.
0.4 1.6138 50.20 21 .infin. 0.01 1.5220 63.00 22 .infin. 0 Distance
to the object = 0 Image height = 0.500
[0066] Table 3 below lists the values of the variables of interest
for Embodiments 1 and 2. TABLE-US-00003 TABLE 3 item legend unit
Embodiment 1 Embodiment 2 image scale factor .beta.o -2.678847
-6.63 focal length of front f1 [mm] 0.765 0.591 lens group focal
length of rear f2 [mm] 3.476 4.557 lens group focal length f [mm]
0.657 0.797 half of field angle wy [deg] 6.141 3.95 exit angle of
chief ray wy' [deg] 13.965 6.02 numerical aperture on NA 0.2184
0.55 the object side stop diameter .PHI.1 [mm] 0.36 0.66 largest
lens diameter .PHI.2 [mm] 1 1.2 pitch P [.mu.m] 4 4 reference
wavelength .lamda. [.mu.m] 0.546 0.546
[0067] Table 4 below lists the Conditions (1)-(6) and the values
corresponding thereto for Embodiments 1 and 2. TABLE-US-00004 TABLE
4 Condition No. Condition Embodiment 1 Embodiment 2 1 1 <
|.beta.o| .ltoreq. 10 2.680 6.630 2 0.9 .ltoreq. |cos wy'/cos wy|
.ltoreq. 1.1 0.976 0.997 3 0.1 .ltoreq. NA .ltoreq. 0.8 0.220 0.550
4 0.1 .ltoreq. |p NA.sup.2/(0.61 .lamda. .beta.o)| .ltoreq. 0.8
0.215 0.544 5 0.2 .ltoreq. .PHI.1/(.PHI.2 f1) .ltoreq. 2 0.471
0.931 6 2 .ltoreq. f2/f1 .ltoreq. 10 4.544 7.711
[0068] A video observation system suitable for in vivo cellular
observation will now be described. As mentioned above, living
tissues of interest often include parenchymal tissues with
transparent epithelial cells overlying the parenchymal tissues. The
following techniques are used in order to create sufficient
contrast between the cell nuclei and other cell portions so as to
enable observation of the epithelial cells within a targeted
observation region with no interference from the underlying
parenchymal tissues. For example, a video observation system
suitable for distinctly observing a layer of cells that have been
stained blue for improved contrast has the following
configuration.
[0069] FIG. 3 shows a configuration for the video observation
system that uses the magnifying image pickup unit of the present
invention. The illumination light supplied by a light source device
11 illuminates an object 12 via a magnifying image pickup unit 13.
The light source device 11 is provided with a wavelength selection
filter 14, which is positioned in the illumination light path, as
required, to produce illumination light having a wavelength profile
suitable for cellular observation. When illumination light having a
visible wavelength range is used to illuminate living tissue, the
shorter wavelengths that correspond to blue reach only the surface
of the living tissue. These wavelengths are useful to obtain
information specific to the epithelial cells of the living tissue.
Light having wavelengths around 500 nm (corresponding to the color
green) reaches only slightly below the surface of living tissue. On
the other hand, light wavelengths corresponding to the color red
reach relatively deep inside living tissue.
[0070] An image is formed by the objective optical system of the
magnifying image pickup unit 13 at the image pickup surface of the
image pickup element. The image pickup unit converts the image into
electrical signals and sends them to an image processing unit 15.
In processing image data of the visible region, green wavelength
components are used to produce the brightness information of the
object. In this way, images are obtained that are similar to those
acquired through the human eye. The image data that are processed
by the image processing unit 15 are displayed on a TV monitor
16.
[0071] When the cells that have been stained blue are illuminated
by white light that equally contains blue, green, and red
components, the portions that have been stained blue appear blue
while unstained portions appear white. FIG. 4 shows a wavelength
profile of a cellular image formed by the magnifying image pickup
unit, with the Y-axis (the ordinate) being the light intensity in
arbitrary units and the X-axis (the abscissa) being the wavelength
in nm. Unstained portions do not absorb specific light wavelengths
and thus provide a nearly flat wavelength profile, as is shown by
the solid lines 17. Portions that have been stained blue absorb red
light and give a wavelength profile with a drop in intensity
primarily for the red component, as shown by the broken lines
18.
[0072] In this way, the contrast between the background (the
unstained portions, shown by the solid lines) and the cell nuclei
and walls (the stained portions, shown by the broken lines) appears
as a difference in light reflection 19 that is due to light
absorption that occurs primarily at the longer wavelengths (i.e.,
the red component) for images obtained using the magnifying image
pickup unit.
[0073] As shown in FIG. 1, the epithelial cells are vertically
layered. When observing the cells in a specific layer, the images
of layers of cells that are not at the depth of interest in the
epithelial cells and the cells of the underlying parenchymal
tissues overlap in the background, thereby reducing the image
contrast. Cell walls are only slightly stained by the staining
processes as compared to cell nuclei. FIG. 6 illustrates the
situation where the images of cells that are not of interest
overlap in the background, and the cell walls are barely
recognizable.
[0074] In order to eliminate unwanted images (i.e., noise)
overlapping in the background, the light that transmits information
about the parenchymal tissues, and the cell layers that are not at
the depth of interest can be cut off in the illumination light path
or in the objective optical system before the light is detected by
the image pickup element. In this embodiment of the video
observation system, a filter for cutting off a specific range of
wavelengths is inserted in the illumination light path in order to
eliminate unnecessary wavelength components from the illumination
light. In this case, unnecessary components are mainly the longer
wavelengths of the visible light range, such as red light. However,
it is not desirable to eliminate all the red light components
because this cuts off the wavelength components that serve to
provide contrast between the portions that have been stained blue
and the unstained portions.
[0075] The present invention uses, for the wavelength selection
filter 14 of the light source, a filter having a spectral
transmittance as shown in FIG. 7. FIG. 7 shows the spectral
transmittance by solid line with the ordinate (i.e., the Y-axis)
being the transmittance and the abscissa (i.e., the X-axis) being
the wavelength in nm. More specifically, among the illumination
light that includes the blue, green, and red wavelength ranges, the
illumination light of the red wavelength range is divided into two
wavelength bands R1 and R2. Among the wavelength bands R1 and R2,
the wavelength band R1 that is nearer the green wavelength range is
cut off by the filter and thus prevented from illuminating the
object. The wavelength band R1 may be, for example, the range 600
nm<.lamda.<700 nm, and the wavelength band R2 may be, for
example, the range 700 nm<.lamda.<800 nm. FIG. 8 shows an
image in which information from regions other than regions at a
desired depth is eliminated.
[0076] Using illumination light with the wavelength band R1 cut off
allows limited light to reach the parenchymal tissues and the cell
layers that are not at the depth of interest, but are within the
depth of field of the objective optical system. Thus, the image
resolution in the depth direction is improved by preventing
overlapping of unwanted images. On the other hand, the light in the
wavelength band R2 contributes to producing contrast between the
cell nuclei that have been stained blue verses the cell walls and
other cell portions that remain relatively unstained. Thus, a clear
image of the cell layer of interest is obtained with the
information from unnecessary depths being eliminated. The
transmittance characteristic shown in FIG. 7 can be realized using
a dichroic filter.
[0077] A contrast medium can be used to enhance only the cell
nuclei. In such a case, the contrast medium that is absorbed by the
cell nuclei has outer electrons that are excited by excitation
light and which emit fluorescent light when they return to the
ground state. This fluorescent light can be observed in order to
accurately identify the cell nuclei. In particular, the video
observation system according to the present invention is useful for
a method where a gene contrast medium, such as a gene marker (for
example GFP) that reacts with light, can be injected into the cells
and the identification of a specific gene that occurs when healthy
cells are transformed into diseased cells such as cancer may be
accomplished.
[0078] In the method above, the gene in a living cell is altered
immediately before the onset of disease and a gene marker, which
takes no action among the normal cells, identifies diseased areas
and emits weak fluorescence in response to excitation light. Thus,
the video observation system used for this observation is desirably
provided with a hypersensitive camera.
[0079] It is preferable that the video observation system for
fluorescent image observation of cell nuclei be used in combination
with a conventional endoscope. FIG. 15 shows an example of such a
combination. The fluorescent image observation system is formed as
a thin endoscope that has an elongated portion 41. A magnifying
image pickup unit 42 of the fluorescent image observation system is
mounted on the distal end of the elongated portion. The reference
numeral 43 denotes a conventional endoscope that has a channel 44
that extends from the distal end to the proximal end (not shown) of
the conventional endoscope. The conventional endoscope also has an
observation window 45 as well as illumination windows 46 and 47 at
its distal end. The channel is also used for inserting treatment
tools. The elongated portion 41 of the fluorescent image
observation system is inserted into the channel 44 from the
proximal end of the conventional endoscope 43 and comes out of the
channel 44 via the opening in the channel. The endoscope
observation system is inserted into a body cavity to be observed.
The conventional endoscope provides images of the elongated portion
41 that protrudes from the channel for guiding the magnifying image
pickup unit 42 to the targeted observation region. The TV monitor
16 (shown in FIG. 3) displays images from the conventional
endoscope as well as fluorescent images from the image pickup unit
simultaneously, thereby providing more precise and accurate
observations.
[0080] When a contrast medium that primarily absorbs light of
wavelengths shorter than 480 nm and emits fluorescent light having
wavelengths longer than 470 nm is used, a preferred wavelength
selection filter 14 for the light source has a spectral
transmittance as shown in FIG. 9. In FIG. 9, the spectral
transmittance is shown by the solid lines with the ordinate
illustrating the transmittance and the abscissa being the
wavelength in nm.
[0081] More specifically, the illumination light of the blue
wavelength range is divided into two wavelength bands B1 and B2
and, among the wavelength bands B1 and B2, the wavelength band B1
that is nearer the green wavelength range is cut off by the filter.
For example, the wavelength band B1 may lie in the range 450
nm<.lamda.<500 nm and the wavelength band B2 may lie in the
range 350 nm<.lamda.<450 nm. In such a case, the objective
optical system may be provided with a filter that transmits light
having wavelengths longer than about 470 nm and cuts off light
having wavelengths shorter than about 470 nm. Thus, fluorescent
images can be observed while the excitation light is cut off. The
light of the wavelength band B1 that includes the excitation and
fluorescent wavelengths is cut off from the illumination light.
This enables clear images using weak fluorescence to be observed
with no interference from the excitation light.
[0082] The wavelength selection filter 14 for the light source
device can be used with a filter that reduces the light intensity
of either the green or red wavelength range. This prevents
unnecessary image noise in the background of fluorescent images of
the cell nuclei and allows a conventional endoscope to produce
conventional observation images of living tissue.
[0083] An illumination method suitable for in vivo cellular
observation is described hereafter with reference to FIG. 13. The
tip of an illumination unit 30 that serves to illuminate an object
and the tip of an objective optical system 31 that serves to form
images on the image pickup surface of an image pickup element using
light from the object are located at the tip 32 of an endoscope.
The central axis L of the illumination field of the illumination
unit 30 is substantially parallel to and shifted from the central
axis F of the field of view of the objective optical system 31 by a
distance d; thus, the center line of the illumination field and the
center line of the objective optical system (i.e., of the
observation field) are directed in substantially the same
direction. The endoscope tip 32 is placed adjacent living tissue in
order to observe the living tissue. The distance between a targeted
region of the living tissue and the endoscope tip is adjusted so
that the targeted region among the epithelial cells and parenchymal
tissues which form the living tissue is "in-focus" (i.e., centrally
located within the depth of field of the objective optical system).
As shown in FIG. 13, the distance between the position of the
epithelial cells 34 and endoscope tip 32 is X1 and the distance
between the position of the parenchymal tissues 33 and the
endoscope tip 32 is X2.
[0084] The conventional endoscope uses the field of view F2 of the
objective optical system to observe the parenchymal tissues at the
distance of X2. The distance d between the central axis L of the
illumination field and the central axis F of the field of view at
the endoscope tip is determined in a manner such that the
illumination field L2, which is positioned in front of the
endoscope tip by a distance X2, includes the field of view F2 in
order to ensure a uniform brightness in the field of view F2. The
distance X2 is several millimeters to several tens of
millimeters.
[0085] On the other hand, the magnifying endoscope uses an
objective optical system having a field of view F1 in order to
observe the epithelial cells at a distance X1. The distance X1 is
within the range of zero to several microns, that is, substantially
zero. Therefore, as shown in FIG. 13, the illumination field L1a,
which is positioned in front of the endoscope tip by the distance
X1, may fail to include the field of view F1 of the magnifying
endoscope even when the distance d between the central axes of the
illumination field L and the field of view F at the endoscope tip
is reduced. Consequently, it is understood that the conventional
illumination method fails to ensure a uniform brightness in the
field of view F1 of the objective optical system. The present
invention provides an illumination method in which the parenchymal
tissues that are positioned still farther in front of the endoscope
tip than the farthest position of the depth of field of the
objective optical system are utilized for uniformly illuminating
the field of view F1 of the objective optical system.
[0086] As shown in FIG. 13, an observation target is at a distance
X1 from the endoscope tip 32 and the parenchymal tissues 33 are at
a distance of X2 from the endoscope tip. The illumination light
emitted from the endoscope tip 32 reaches the parenchymal tissues
via the illumination field L1a. The parenchymal tissues 33 serve as
reflecting and scattering surfaces and thus scatter the
illumination light. It is assumed that the illumination light
emitted from the endoscope tip 32 has a Gaussian light distribution
profile, and the effective light distribution angle of illumination
will herein be defined as a light distribution angle .omega. that
provides a light intensity that is 1/e times the light intensity
on-axis, where e is the base of the natural logarithm.
[0087] As shown in FIG. 13, the illumination light emitted from the
endoscope tip 32 is transmitted through the living tissues at the
light distribution angle .omega.1' before it reaches the
parenchymal tissues 33. After being reflected and scattered by the
parenchymal tissues, the illumination light is emitted at the light
distribution angle .omega.2' and thereafter it reaches the
epithelial cells at the in-focus region 34 and forms the
illumination field L1b at a distance X1 from the endoscope tip that
includes the field of view F1 of the objective optical system.
Consequently, uniform brightness is ensured in the field of view F1
of the objective optical system. Thus, an object placed in contact
with a distal end of the observation unit so that a light source
that does not directly illuminate an observation field of view
illuminates an area of tissue outside the observation field of
view, and the illuminated tissue scatters light from the light
source so as to illuminate the observation field of view. The
observation unit is then used to observe an image of the
observation field with a scale factor larger than 1.
[0088] The light distribution angles .omega.1' and .omega.2' are
light distribution angles within living tissues. The following
Equations (B) and (C) are used to convert these light distribution
angles to the equivalent light distribution angles .omega.1,
.omega.2 in air. sin .omega.1=1.33sin .omega.1' Equation (B) sin
.omega.2=1.33sin .omega.2' Equation (C)
[0089] The parenchymal tissues are outside the depth of field of
the objective optical system, therefore, the light reflected and
scattered by the parenchymal tissues is not imaged and thus merely
the illumination effect on the epithelial cells is obtained. It is
preferred that the distance d at the endoscope tip between the
central axes of the illumination field L and the field of view F
satisfies the following Condition (7): 1.ltoreq.log(d/(X1tan
.omega.)).ltoreq.3 Condition (7) where
[0090] d is the distance between the central axis of the
illumination field and the central axis of the field of view of the
objective optical system,
[0091] X1 is the distance between the leading surface of the
endoscope (i.e., the endoscope tip) and the in-focus point of the
objective optical system, and
[0092] .omega. is the light distribution angle that provides a
light intensity that is lie times the light intensity on-axis,
where e is the base of the natural logarithm.
[0093] When the upper limit of Condition (7) is not satisfied,
uniform brightness will not be ensured in the field of view of the
objective optical system. When the lower limit of Condition (7) is
not satisfied, it will be difficult to locate the tips of the
illumination unit and the objective optical system at the endoscope
tip with the outer diameter of the endoscope tip being maintained
small.
[0094] In addition, the following Condition (8) is preferably
satisfied: 5.ltoreq.d/(X2tan .omega.).ltoreq.30 Condition (8)
where
[0095] d and .omega. are as defined above, and
[0096] X2 is the distance between the leading surface of the
endoscope (i.e., the endoscope tip) and the reflecting and
scattering surfaces, such as the parenchymal tissues.
[0097] When the upper and lower limits of Condition (8) are not
satisfied, uniform brightness will not be ensured in the field of
view of the objective optical system.
[0098] It is also preferred that the following Condition (9) be
satisfied: 0.5.ltoreq.log(X2/X1) Condition (9) where
[0099] X2 and X1 are as defined above.
[0100] When Condition (9) is not satisfied, the parenchymal tissues
are imaged in the field of view, deteriorating the image
quality.
[0101] Table 5 below lists various values of X2, X1, d and .omega.
pertaining to Embodiments 1 and 2 of the present invention.
TABLE-US-00005 TABLE 5 item legend unit Embodiment 1 Embodiment 2
scatterer distance X2 [mm] 0.1 0.1 objective in-focus X1 [mm] 0.015
0.002 distance illumination parallax d [mm] 0.8 1 illumination
.omega. [deg] 35 35 distribution angle
[0102] Table 6 below lists the Conditions (7)-(9) and the value of
each for Embodiments 1 and 2. TABLE-US-00006 TABLE 6 Condition No.
Condition Embodiment 1 Embodiment 2 7 1 .ltoreq. log (d/(X1 1.88
2.85 tan .omega.)) .ltoreq. 3 8 5 .ltoreq. d/(X2 tan .omega.)
.ltoreq. 30 11.4 14.3 9 0.5 .ltoreq. log (X2/X1) 0.82 1.7
[0103] The method for diagnosing the presence/absence of abnormal
cells (i.e., whether cells are cancerous or not) from magnified
cell images will now be described.
[0104] FIG. 5 shows stained cells that are magnified and observed
according to Embodiment 1. The image pickup unit used in the
magnifying endoscope observation system is set for a scale factor
that allows several tens to several hundreds of nuclei to be
observed on a monitor. For example, several tens to several
hundreds of cell nuclei are displayed on a monitor and the cell
density in the observation field of view can be evaluated based on
the distance between cell nuclei in order to diagnose the
presence/absence of abnormal cells. The cell density can be
compared with normal samples and statistically analyzed.
[0105] The magnifying endoscope observation system above is
specified for a resolution and a magnification sufficient for
nucleic observation. With the observation scale factor further
increased, the image pickup unit displays several cell nuclei on a
monitor and the number of cell nuclei in a unit area is translated
to the cell size, or the cell nuclei are evaluated for shape, in
order to diagnose the presence/absence of abnormal cells. For
example, cancerous cells present particular characteristics such as
increased size and irregular shapes. Thus, the size and shape of
cell nuclei can be evaluated in order to diagnose cancerous
cells.
[0106] FIG. 8 shows cell nuclei and cell walls that are magnified
and observed according to Embodiment 2 of the present invention.
The magnifying endoscope observation system is set for a scale
factor that allows several nuclei to be observed on a monitor. The
ratio of the area S' of the cell nuclei divided by the area S
within the cell walls in the field of view is herein defined as the
"occupancy" of the nuclei in the cells, and is used to diagnose the
presence/absence of abnormal cells. For this analysis, the
magnifying endoscope observation system is specified so as to have
a resolution and contrast sufficient for observation of both cell
nuclei and cell walls.
[0107] FIG. 14 is a flow chart of a series of procedures for the in
vivo cellular observation described above. As noted in the drawing,
there is a pre-treatment stage, followed by a cell visualization
stage, followed by a stage of abnormal cell diagnosis. During
pre-treatment, the following steps are performed: detect a
suspicious region by using a conventional endoscope; deliver a
coloring agent using a conventional endoscope; selectively stain
cell parts due to differences in time for cells to intake or
excrete the coloring agent; and guide a magnifying endoscope to the
suspicious region and contact the tip thereof to the region for
observation. During cell visualization the following step is
performed: emit illumination light having selected wavelengths; the
illumination light including wavelength band T2, with the
wavelength band T1 being nearer the green wavelength range than is
the wavelength band T2, for visualizing cells due to differences in
absorbency, and wavelength band T1 for cutting off data at depths
not targeted. During abnormal cell diagnosis, the following steps
are performed: display several tens to several hundreds of cell
nuclei; and, display several cell nuclei and the cell walls. Note
that in CELL VISUALIZATION of FIG. 14, the second and third boxes
are not separate steps; instead, these boxes merely indicate the
details of the selected wavelengths.
[0108] The system described above in which specific wavelength
bands are used from a white light source provides excellent
flexibility where plural wavelength properties are selectively used
in the illumination light, depending on the observation target and
the selected coloring agent. On the other hand, where the
wavelength property of the illumination light is predetermined, a
single color illumination light, for example, can be used to
further simplify the configuration.
[0109] An endoscope tip part of an endoscope imaging system
specified for observation with specific wavelengths of light is
illustrated in FIGS. 10(a) and 10(b), with FIG. 10(a) being a side
cross section and FIG. 10(b) being an end view. The image pickup
unit according to the present invention has a small observation
distance and therefore, the light source can be formed of LEDs 20
that emit a single color at low power. The LEDs 20 can be mounted
in the endoscope body. Furthermore, when LEDs are installed at the
tip of the endoscope, a light transmission optical fiber can be
eliminated. Because aberration correction is necessary for only a
single color when an LED light source is used, the fluorescent
image observation system 21 can be provided with a simplified
optical system that includes, for example, a single aspherical lens
23 positioned in front of a stop 24, and an image pickup element 25
positioned after the stop 24 at the image surface. Alternatively,
plural spherical lenses (not shown) can be used in lieu of using a
single aspherical lens.
[0110] An image pickup element that has been made compact and
simplified as described above provides more freedom in mounting
such an image pickup unit on a medical device such as an endoscope.
For example, an image pickup unit may be combined with a treatment
tool such as a catheter or laser probe that uses a flexible,
insertable device. Or, an image pickup unit may be combined with a
non-flexible treatment tool by making the image pickup unit compact
and with a shape such as a pen or capsule by using wireless
transmission of image data.
[0111] The invention being thus described, it will be obvious that
the same may be varied in many ways. Such variations are not to be
regarded as a departure from the spirit and scope of the invention.
Rather, the scope of the invention shall be defined as set forth in
the following claims and their legal equivalents. All such
modifications as would be obvious to one skilled in the art are
intended to be included within the scope of the following
claims.
* * * * *