Title: Measuring apparatus
Abstract: A measuring apparatus for measuring the state of attenuated total reflection over time for a single measuring unit without being affected by the change in measuring conditions arising from replacement and resetting of the sample. A light beam is entered into the interface between a dielectric block and a metal film having a dielectric film thereon at various incident angles within the angle range that creates two or more dark lines due to attenuated total reflections, and the variation in the positions of other dark lines are measured with reference to the dark line having the least positional variation among them.
Patent Number: 6,947,145 Issued on 09/20/2005 to Naya
| Inventors:
|
Naya; Masayuki (Kaisei-machi, JP)
|
| Assignee:
|
Fuji Photo Film Co., Ltd. (Kanagawa-ken, JP)
|
| Appl. No.:
|
630713 |
| Filed:
|
July 31, 2003 |
Foreign Application Priority Data
| Jul 31, 2002[JP] | 2002-223231 |
| Current U.S. Class: |
356/445 |
| Intern'l Class: |
G01N 021/17 |
| Field of Search: |
356/445-448
|
References Cited [Referenced By]
U.S. Patent Documents
| 5485277 | Jan., 1996 | Foster.
| |
| 2002/0140938 | Oct., 2002 | Naya et al.
| |
| 2003/0075697 | Apr., 2003 | Ohtsuka et al.
| |
| 2003/0156292 | Aug., 2003 | Naya.
| |
| Foreign Patent Documents |
| 6-167443 | Jun., 1994 | JP.
| |
| 2003/-227792 | Aug., 2003 | JP.
| |
Primary Examiner: Stafira; Michael P.
Attorney, Agent or Firm: Sughrue Mion, PLLC
Claims
1. A measuring apparatus comprising:
a measuring unit having a transparent dielectric block, a metal film formed on
one of the surfaces of said dielectric block, and a transparent dielectric film
formed on said metal film;
a light beam entering means for entering a light beam into said dielectric block
at various incident angles within an angle range that satisfies the conditions
of total reflection at the interface between said dielectric block and said metal
film, and creates two or more dark lines due to attenuated total reflections in
a light beam totally reflected at said interface;
a light detecting means for receiving said light beam totally reflected at said
interface, and detecting positions on a light receiving surface of said two or
more dark lines contained therein; and
a calculation means for calculating a variation in each of said positions of
said two or more dark lines on said light receiving surface arising from a change
in the dielectric constant of a substance placed on said transparent dielectric
film with reference to one of said two or more dark lines having the least positional
variation on said light receiving surface among said two or more dark lines, based
on the output of said light detecting means.
2. A measuring apparatus according to claim 1, wherein said dark line having
the least positional variation is a dark line created by a light component of said
light beam having the largest incident angle at said interface among said two or
more dark lines.
3. A measuring apparatus according to claim 1, wherein said measuring unit further
comprises a sensing material fixed on said dielectric film, and said change in
the dielectric constant is a change in said dielectric constant arising from a
reaction when a test substance containing a material that reacts to said sensing
material is brought into contact with said sensing material.
4. A measuring apparatus according to claim 2, wherein said measuring unit further
comprises a sensing material fixed on said dielectric film, and said change in
the dielectric constant is a change in said dielectric constant arising from a
reaction when a test substance containing a material that reacts to said sensing
material is brought into contact with said sensing material.
5. A measuring apparatus according to claim 1, wherein said metal film has a
thickness of 10 nm to 70 nm, and said transparent dielectric film has a thickness
of 100 nm to 2000 nm.
6. A measuring apparatus according to claim 2, wherein said metal film has a
thickness of 10 nm to 70 nm, and said transparent dielectric film has a thickness
of 100 nm to 2000 nm.
7. A measuring apparatus according to claim 3, wherein said metal film has a
thickness of 10 nm to 70 nm, and said transparent dielectric film has a thickness
of 100 nm to 2000 nm.
8. A measuring apparatus according to claim 4, wherein said metal film has a
thickness of 10 nm to 70 nm, and said transparent dielectric film has a thickness
of 100 nm to 2000 nm.
9. A measuring apparatus according to claim 1, wherein said transparent dielectric
film is made of SiO
2, a glass, or plastic material.
10. A measuring apparatus according to claim 2, wherein said transparent dielectric
film is made of SiO
2, a glass, or plastic material.
11. A measuring apparatus according to claim 3, wherein said transparent dielectric
film is made of SiO
2, a glass, or plastic material.
12. A measuring apparatus according to claim 4, wherein said transparent dielectric
film is made of SiO
2, a glass, or plastic material.
13. A measuring apparatus according to claim 5, wherein said transparent dielectric
film is made of SiO
2, a glass, or plastic material.
14. A measuring apparatus according to claim 6, wherein said transparent dielectric
film is made of SiO
2, a glass, or plastic material.
15. A measuring apparatus according to claim 7, wherein said transparent dielectric
film is made of SiO
2, a glass, or plastic material.
16. A measuring apparatus according to claim 8, wherein said transparent dielectric
film is made of SiO
2, a glass, or plastic material.
Description
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates to a measuring apparatus for sample analysis using
an evanescent wave generated when a light beam is totally reflected.
2. Description of the Related Art
In a metal, free electrons oscillate as a group and a compressional wave called
a plasma wave is generated. The quantized compressional waves generated on the
surface of a metal are known as surface plasmons.
Various surface plasomon sensors have been proposed for quantitative analysis
of a substance in a sample by applying the phenomenon that the surface plasmons
are excited by a light wave. Among these sensors, the one that uses a system called
Kretschmann geometry is particularly well-known as described, for example, in Japanese
Unexamined Patent Publication No.6 (1994)-167443.
Basically, the surface plasmon sensor that uses the system described above
comprises, for example, a dielectric block shaped like a prism; a metal film formed
on one of the surfaces of the dielectric block and brought into contact with a
sample; a light source for generating a light beam; an optical system for entering
the light beam into the dielectric block at various angles to satisfy the conditions
of total reflection at the interface between the dielectric block and the metal
film, and to cause attenuated total reflection by surface plasmon resonance; and
a light detecting means for detecting the state of surface plasmon resonance or
attenuated total reflection by measuring the intensity of the light beam totally
reflected at the interface.
In order to obtain a light beam having the various incident angles described
above,
a comparatively narrow light beam may be entered into the interface by changing
its incident angle, or a comparatively wide light beam may be entered thereto as
a converging or diverging light beam to include light components that are incident
on the interface at various angles. In the first case, the reflected light beam
that changes its reflection angle in accordance with its incident angle may be
detected by a small light detector that moves in synchronization with the change
in the reflection angle, or by an area sensor that extends along the changing direction
of the reflection angle. In the latter case, the reflected light beam may be detected
by an area sensor that extends along the direction where all of the light components
of the light beam reflected at various angles may be captured.
When a light beam enters the metal film of a surface plasom sensor configured
in the aforementioned manner at a certain incident angle θ
sp which
is greater than the total reflection angle, an evanescent wave having a distributed
electric field in the sample in contact with the metal film is generated, and thereby
surface plasoms are excited at the interface between the metal film and the sample.
When the wave-number matching is achieved, in which the wave-number vector of the
evanescent light matches the wave-number vector of the surface plasmons, the evanescent
light and the surface plasmons go into the sate of resonance, and the intensity
of the light totally reflected at the interface between the dielectric block and
the metal film drops sharply, because the light energy is transferred to the surface
plasmons. This drop in the intensity of light is generally detected as a dark line
by the light detecting means described above.
The resonance described above occurs only when a p-polarized light beam enters
the metal film. Accordingly, arrangements need to be made in advance so that the
light beam enters the metal film in p-polarized mode, or only p-polarized light
components are detected by the light detecting means.
When the wave-number of the surface plasmons is determined from the incident
angle θ
sp that causes attenuated total reflection (ATR), the dielectric
constant of the sample may be obtained. More specifically, the following relationship
may be derived, assuming that Ksp as the wave-number of surface plasmons, ω
as the angular frequency of the surface plasmons, c as speed of light in vacuum,
ε
m as the dielectric constant of the metal, and ε
s
as the dielectric constant of the sample.
##EQU1##
When the dielectric constant ε
s is determined, the density
of a particular substance in the sample may be obtained based on a predefined calibration
curve, and the like. That is, by determining θ
sp that causes the
drop in the reflected intensity of light described above, the dielectric constant
of the sample or characteristics related to the refractive index of the sample
may be obtained.
The leakage mode sensor described, for example, on pages 21 to 23, and 26 to
27 of "BUNKOH KENKYU", Vol. 47, No.1 (1998) is also known as a similar sensor that
uses attenuated total reflection (ATR). Basically, the leakage mode sensor comprises,
for example, a dielectric block shaped like a prism; a cladding layer formed on
one of the surfaces of the dielectric block; an optical guiding layer formed on
the cladding layer and brought into contact with the sample; a light source for
generating a light beam; an optical system for entering the light beam into the
dielectric block at various angles to satisfy the conditions of total reflection
at the interface between the dielectric block and the cladding layer, and to cause
attenuated total reflection by the excitation of guided mode in the optical guiding
layer; and a light detecting means for detecting the state of excitation of guided
mode or attenuated total reflection by measuring the intensity of the light beam
totally reflected at the interface.
When a light beam is incident on the cladding layer through the dielectric block
of a leakage mode sensor configured in the aforementioned manner at a certain incident
angle which is equal to or greater than the total reflection angle, certain light
components of the light beam having particular wave-numbers and incident angles
pass through the cladding layer, and propagate along the optical guiding layer
in guided mode. When the guided mode is excited in this manner, attenuated total
reflection occurs, in which the intensity of the light totally reflected at the
interface described above drops sharply, because most of the light components of
the light beam are contained in the optical guiding layer. The wave-number of the
guided light is dependent on the refractive index of the sample placed on the optical
guiding layer, so that the refractive index of the sample and other characteristics
related thereto may be analyzed by determining the particular incident angle that
causes the attenuated total reflection described above.
Various types of measuring apparatuses are available that utilize attenuated
total reflection of a surface plasmon sensor, leakage mode sensor, or the like,
such as an apparatus that enters a light beam containing a plurality of light components
of different wavelength into the interface, and detects the level of attenuated
total reflection for each wavelength, or an apparatus that splits up a portion
of the light beam to be entered into the interface before entering and mixes up
the split-up light beam with the light beam reflected at the interface to interfere
with each other, and measures the state of the interference, as well as an apparatus
that measures a particular incident angle that causes attenuated total reflection
described above, in the process of analyzing characteristics of a subject under
test by entering light to the interface with an angle that satisfies the conditions
of total reflection, and measuring changes in the state of the light totally reflected
at the interface due to the evanescent wave generated by the light entered into
the interface.
In the conventional surface plasmon sensor, or leakage mode sensor described
above,
a sample is sometimes replaced together with the dielectric block in order to efficiently
conduct measurement for a plurality of samples, when the sample (same measuring
unit) needs to be measured a plurality of times at intervals to analyze changes
in the state of the sample over time. In this case, when the sample is reset on
the measuring apparatus for measurement after replacement, a difference (in inclination)
may arise between the initial baseline (interface described above) and the latter
baseline. If this difference in inclination between the two baselines is a difference
in the vertical inclination that changes the incident angle of the light beam entering
at various incident angles, then the reflection angle of the reflected light beam
is also deviated, thereby the accuracy of the measurement is lost.
Even when a sample replacement does not take place, changes in the inclination
of the baseline may occur subtly by vibrations, or the like, when a plurality of
measuring units is moved or rotated on a support or rotating platform. In such
a case, changes in the inclination of the baseline developed during a plurality
of measurements cause measurement errors.
SUMMARY OF THE INVENTION
The present invention has been developed in recognition of the circumstance described
above and it is an object of the present invention to provide a high-accurate measuring
apparatus by preventing measurement errors caused by changes in the inclination
of the interface where a light beam is totally reflected, in conducting measurement
a plurality of times for a single measuring unit.
The measuring apparatus according to the present invention comprises:
a measuring unit having a transparent dielectric block, a metal film formed on
one of the surfaces of said dielectric block, and a transparent dielectric film
formed on said metal film;
a light beam entering means for entering a light beam into said dielectric block
at various incident angles within an angle range that satisfies the conditions
of total reflection at the interface between said dielectric block and said metal
film, and creates two or more dark lines due to attenuated total reflections in
a light beam totally reflected at said interface;
a light detecting means for receiving said light beam totally reflected at said
interface, and detecting positions on a light receiving surface of said two or
more dark lines contained therein; and
a calculation means for calculating a variation in each of said positions of
said
two or more dark lines on said light receiving surface arising from a change in
the dielectric constant of a substance placed on said transparent dielectric film
with reference to one of said two or more dark lines having the least positional
variation on the light receiving surface among said two or more dark lines, based
on the output of said light detecting means.
At least one of the "two or more dark lines" is a dark line created by the attenuated
total reflection due to surface plasmon resonance occurred on the interface between
the dielectric block and the metal film, and others are those created by the attenuated
total reflection due to excitation of light-guided mode.
More specifically, the measuring apparatus according to the present invention
causes attenuated total reflections by the excitation of surface plasmons and light-guided
mode simultaneously, which are respectively the feature of the surface plasmon
sensor and the leakage mode sensor as described above, and detects the dark lines
created by these attenuated total reflections. The applicant has found that providing
a transparent dielectric film on the metal film of the measuring unit causes the
change in the dielectric constant of a substance (sample) placed thereon to have
an impact only on the wave-number that induces light-guided mode, and substantially
not on the wave-number that induces surface plasmons, creating a stable dark line
that remains at substantially the same position on the light receiving surface.
The measuring apparatus according to the present invention opened a way for accurate
measurement by preventing measurement errors caused by, for example, changes in
the inclination of the interface, and the like by incorporating the finding described
above and by measuring a variation in each of the positions of the dark lines on
the light receiving surface using the aforementioned stable dark line as the reference.
In addition, the measuring apparatus according to the present invention enters
a light beam into the interface in p-polarized mode with respect to the interface
in order to achieve surface plasmon resonance.
Further, "the position on the light receiving surface" described above corresponds
to the incident angle of the light beam at the interface (the reflection angle
at the interface), and detecting the position of the dark line on the light receiving
surface is equivalent to detecting the incident angle of the light beam at the interface.
As for the reference dark line described above, a dark line created by a light
component having the largest incident angle at the interface may be used among
the two or more dark lines described above.
A sensing material may be fixed on the dielectric film of the measuring unit
described
above, and the change in the dielectric constant described above may be the change
in the dielectric constant arising from a reaction when a test substance containing
a material that reacts to the sensing material is brought into contact with the
sensing material. That is, in this case, both the sensing material and material
that reacts to the sensing material are the intended samples for the measurement.
Preferably, the thickness of the metal film is 10 nm to 70 nm, and the
thickness of the transparent dielectric film is 100 nm and 2000 nm
Preferably, the transparent dielectric film is made of SiO
2,
a glass, or plastic material.
Further, the dielectric block described above may be a dielectric block
formed as a single block having the entrance and outgoing surfaces for the light
beam, and a surface the metal film is formed thereon, or it may comprise a component
having the entrance and outgoing surfaces for the light beam, and a component having
a surface the metal film is formed thereon, connected to each other through a refractive
index matching means.
As for the light detecting means for the measuring apparatuses described above,
a photodiode array, CCD, or the like may be preferably used.
The measuring apparatus according to the present invention comprises a measuring
unit having a transparent dielectric block, a metal film formed on one of the surfaces
of the dielectric block, and a transparent dielectric film formed on the metal
film, so that it may create two or more dark lines due to attenuated total reflections
when a light beam is entered into the interface between the dielectric block and
the metal film at various incident angles through the dielectric block.
Further, the measuring apparatus according to the present invention further
comprises a light beam entering means for entering a light beam into the interface
at various incident angles within the angle range that creates two or more dark
lines, and a light detecting means for detecting the positions on the light receiving
surface of the two or more dark lines contained in the light beam totally reflected
at the interface, and a calculation means for calculating a variation in each of
the positions of the two or more dark lines on the light receiving surface arising
from a change in the dielectric constant of a substance placed on the transparent
dielectric film with reference to the dark line having the least positional variation
on the light receiving surface among the two or more dark lines, based on the output
of the light detecting means, so that the measuring apparatus may obtain a measurement
result that cancels out errors arising from the change in the inclination of the
interface that vary the incident angle of the light beam at the interface, and
the like.
When the measuring apparatus according to the present invention further comprises
a sensing material fixed on the dielectric film of the measuring unit described
above, and adapted to detect the change in the dielectric constant when a test
substance containing a material that reacts to the sensing material is brought
into contact with the sensing material, it maybe used for the measurement of antigen-antibody
reactions, and the like.
Selection of the thickness of the metal film and the transparent dielectric
film in the ranges from 10 nm to 70 nm, and from 100 nm to 2000 nm respectively
may create two dark lines due to attenuated total reflections, in which case a
signal of higher signal-to-noise ratio may be obtained compared with the case in
which three or more dark lines are created.
BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1 is a side view of a measuring apparatus according to a first embodiment
of the present invention.
FIG. 2 is a drawing illustrating a relationship between the incident angle of
the light beam entered and the reflected intensity of the light beam detected by
the measuring apparatus shown in the FIG. 1.
FIG. 3 is a side view of a measuring apparatus according to a second embodiment
of the present invention.
DESCRIPTION OF THE PREFERRED EMBODIMENTS
Hereinafter, the preferred embodiments of the present invention will
be described in detail with reference to the accompanying drawings.
FIG. 1 is a side view of a measuring apparatus according to a first embodiment
of the present invention.
As shown in the FIG. 1, the measuring apparatus according to the first embodiment
has a measuring chip
10 as a measuring unit. The measuring chip
10
has a test substance holding section
13 made of, for example, transparent
resin, or the like shaped like a frustum of an inverted quadrangular pyramid, and
has a circular-section test substance holding hole
13a on the upper
side. The test substance holding hole
13a is clad with a metal film
12 made of Au on the bottom (a surface
11a of a dielectric
block
11, which will be described later), and a transparent dielectric film
14 made of SiO
2 formed thereon, and a sample solution
15
is held on the transparent dielectric film
14. The lower part of the test
substance holding section
13 of the measuring unit
10 is a dielectric
block
11, and two surfaces facing to each other of the four side surfaces
of the dielectric block are used as a light entrance surface
11b and
a light outgoing surface
11c respectively. That is, the dielectric
block
11 is formed as a single block having the light entrance surface
11b,
the light outgoing surface
11c, and a surface the thin film layer
(metal film)
12 is formed thereon. The dielectric block according to the
first embodiment has a sensing material
30 fixed on the dielectric film
14, which will be described later.
The thickness of the metal film is 10 nm to 70 nm, and the thickness of the transparent
dielectric film is 100 nm to 2000 nm.
Each of the measuring chips
10 is fitted firmly into each of a plurality
of unit holding holes
31a created, for example, through a turntable
31. The turntable
31 rotates intermittently by a certain predefined
angle at a time after the measuring chips
10 are attached thereto in this
manner, and the sample solution
15 is dropped and held in the test substance
holding section
13 of the measuring chip
10 that stopped at a prescribed
position. Thereafter, the turntable
31 further rotates by a certain predefined
angle to move the measuring chip
10 to a position shown in FIG. 1, and stops there.
In addition to the measuring chip
10, which is the measuring unit described
above, the measuring apparatus according to the first embodiment further comprises
a light beam entering means for entering a light beam L from the entrance surface
11b of the dielectric block
11 to an interface
11a
between the dielectric block
11 and the metal film
12 at various
incident angles; a collimator lens
16 for collimating the light beam L totally
reflected at the interface
11a; a light beam detecting means
17
for detecting the collimated light beam L; a signal processing section
20
having a calculation means
20a; and a display section
21 connected
to the signal processing section
20.
The light beam entering means
1 comprises a light source
2 made
of a semiconductor laser, or the like for generating the light beam L, a collimator
lens
3 for collimating the light beam L emitted from the light source
2
in diverging mode, and a converging lens
4 that converges the collimated
light beam L at the interface
11a.
As shown in FIG. 1, the light beam L emitted from the light source
2 in
diverging mode is converged at the interface
11a between the dielectric
block
11 and the metal film
12 by the operation of lenses
3
and
4. Accordingly, the light beam L contains light components having various
incident angles (from θ
1 to θ
2) at the interface
11a,
which are equal to or greater than the total reflection angle. The light beam L
is totally reflected at the interface
11a, and the reflected light
beam L, therefore, contains light components having various reflection angles.
The light beam L is entered into the interface
11a in p-polarized
mode with respect to the interface
11a. In order to obtain a p-polarized
light beam L with respect to the interface
11a, the light source
2 may be prearranged so that the direction of the polarization becomes the
prescribed direction. A wavelength plate or polarization plate may be used as an
alternative means for controlling the direction of the polarization of the light
beam L. Further, the light beam entering means
1 may be configured to enter
the light beam L into the interface
11a in defocused mode. By doing
so, measurement errors in the measurement of the positions of the dark lines described
above are averaged, and hence the accuracy of the measurement is enhanced.
The light beam L totally reflected at the interface
11a and collimated
by the collimator lens
16 is detected by the light detecting means
17.
The light detecting means
17 of the measuring apparatus according to the
first embodiment is a CCD line sensor, which is disposed so that its longitudinal
direction becomes substantially perpendicular to the traveling direction of the
collimated light beam L in the plane of FIG.
1. Accordingly, the light components
of the light beam L totally reflected at the interface
11a at various
angles are received at the respective longitudinal positions of the line sensor.
That is, the position of each of the light components of the light beam L on the
light receiving surface of the line sensor corresponds uniquely to the reflection
angle of each of light components at the interface
11a (i.e., incident
angle at the interface). As for the light detecting means
17, a photodiode
array made of a plurality of photodiodes arranged in the longitudinal direction
of the line sensor may be used, as well as a CCD line sensor.
FIG. 2 shows the relationship between the incident angle θ of the light
beam L totally reflected at the interface
11a and the position on
the light receiving surface of the light detector with its intensity I.
As shown in FIG. 2, when a light beam L is entered into the measuring chip
10
of the measuring apparatus according to the first embodiment, that is, a measuring
chip
10 comprising a metal film
12, and a transparent dielectric
film
14 formed thereon, at incident angles that satisfy the conditions of
total reflection at the interface
11a between the dielectric block
11 and the metal film
12, a plurality of dark lines, S
1 and
S
2, is created due to attenuated total reflections at a plurality of incident
angles within the angle range that satisfies the conditions of total reflection.
One of the dark lines remains substantially unaffected by the change in the dielectric
constant of a sample placed on the transparent dielectric film, while the other
dark line moves widely by the change in the dielectric constant of the sample.
The light component entered the interface
11a at a particular incident
angle θ
S1 excites surface plasmons at the interface between the
metal film
12 and the transparent dielectric film
14, so that the
reflected intensity I of the light component drops sharply (dark line S
1
in FIG.
2). In addition, the light component entered the interface
11a
at another particular incident angle θ
S2 excites light-guided
mode in the transparent dielectric film
14, so that the reflected intensity
I of the light component also drops sharply (dark line S
2 in FIG.
2).
The wave-number of guided light in the transparent dielectric film
14 is
heavily dependent on the dielectric constant of the sample placed on the dielectric
film
14, and if the dielectric constant changes, the incident angle that
creates dark line S
2 due to attenuated total reflection caused by the excitation
of light-guided mode changes, for example, as shown in the dotted line in FIG.
2. That is, the dark line S
2 appeared at the incident angle θ
S2
now appears, for example, at the incident angle θ
S2′.
On the other hand, the dark line S
1 due to attenuated total reflection caused
by the excitation of surface plasmons is dependent on the dielectric constant of
the dielectric film
14 on the metal film
12, which is not affected
by the change in the dielectric constant of the sample, so that the incident angle
θ
S1, that creates the dark line S
1 remains substantially unchanged.
FIG. 2 shows the case in which two dark lines are created, but the light-guided
mode may couple to a plurality of light components having different incident angles.
In that case, a plurality of dark lines that move widely is created by the change
in the dielectric constant of the sample placed on the transparent dielectric film
14. However, a signal of highest signal-to-noise ratio may be obtained under
the condition in which two dark lines are created as shown in FIG.
2. Two
dark lines, one by the excitation of surface plasmons, and the other by the excitation
of light-guided mode, may be obtained by selecting an appropriate thickness for
the metal film and the dielectric film within the respective ranges described above.
The measuring apparatus according to the present invention makes use of the fact
that the position of the dark line S
1 (an incident angle creating the dark
line S
1) on the light receiving surface created by the attenuated total
reflection due to excitation of surface plasmons remains substantially unaffected
by the change in the dielectric constant of the sample, and measures the change
in the dielectric constant of a sample by obtaining a variation in the position
of the dark line S
2 created by the attenuated total reflection due to excitation
of light-guided mode with reference to the dark line S
1.
As described above, the incident angle of the light beam L at the interface corresponds
uniquely to the position on the light receiving surface of the light detector,
so that the change in the incident angles creating the dark lines S
1 and
S
2 may be obtained by detecting the variation in the positions P
1
and P
2 of the dark lines S
1 and S
2 on the light receiving
surface. That is, the change in the incident angles creating the dark lines S
1
and S
2 indicates the change in the dielectric constant (refractive index)
of a sample, so that the change in the dielectric constant of a sample over time
may be obtained by detecting the variation in the positions of the dark lines S
1
and S
2 over time on the light receiving surface of the light detector.
The calculation means
20a calculates the distance ΔP between
a position P
1 of the dark line S
1 and a position P
2 of the
dark line S
2 at each measuring time (ΔP=P
2(t)-P
1(t)).
The signal processing section
20 obtains the change in the dielectric constant
(refractive index) of the sample based on the variation in the ΔP over time,
and the measurement result is displayed on the display
21.
The measuring apparatus according to the first embodiment has a sensing material
fixed on the transparent dielectric film
14, which combines with a particular
substance contained in a sample solution, and the dielectric constant of the sensing
material
30 changes in accordance with the combined state, so that the change
in the combined state may be studied by continuously measuring the distance between
the two dark lines S
1 and S
2. That is, both the sample solution and
the sensing material
30 are the intended samples for analysis. A combination
of such particular substance and the sensing material
30 includes, for example,
a combination of an antigen and antibody.
In addition to calculating the distance ΔP(t) between the two dark lines
at predefined time intervals, the difference between the initial difference ΔP(0)
and the difference ΔP(t) measured after a predetermined time from the initial
measurement (ΔP(t)-ΔP(0)) may be calculated in order to study the change
in the combined state of a particular substance and the sensing material
30
over time.
In measuring the change in the combined state of the sensing material and a particular
substance over time as described above, a single measuring chip is measured a plurality
of times at predefined time intervals. When such measurement is conducted, the
measuring chip
10 is removed from the turntable
31 after it is measured,
and the next measuring chip that holds another sample is mounted on the turntable
31 and measured, then the initial chip
10 is reset thereon for the
next measurement after a predetermined time.
In resetting the measuring chip
10, a difference in the measuring conditions
from the previous measurement may occur. That is, the change in the vertical inclination
of the interface
11a that changes the incident angle of the light
beam L at the interface
11a may occur, causing a deviation in the
positions of the dark lines. As described above, the measuring apparatus according
to the first embodiment, however, always detects two or more dark lines and calculates
the variation in the positions of the dark lines with reference to the dark line
S
1 which is not affected by the change in the dielectric constant of a sample,
so that it may obtain a measurement result that cancels out the influence of the
change, if any, in the inclination of the interface.
As for the cause of the change in the inclination of the interface at each measurement
for a plurality of measurements for a single measuring chip includes but not limited
to the vibration of the support when it is rotated or moved, and relocation of
the support, light source, light detector, and the like, as well as resetting of
the measuring chip described above.
Further, in measuring the state of attenuated total reflection of a sample,
in which the state of attenuated total reflection of the measuring chip
10
itself or the measuring chip
10 that holds only a solvent for the sample
is measured before dispensing the sample solution
15, and the bulk effect
of the measuring chip
10 (and solvent) is subtracted from the measurement
result obtained after dispensing the sample solution, if a change in the inclination
of the interface is developed before and after the dispensing of the sample solution,
the reliability of the measurement result is lost.
The measuring apparatus according to the first embodiment of the present invention
always detects two or more dark lines and calculates the variation in the positions
of the dark lines with reference to the dark line S
1 which is not affected
by the change in the dielectric constant of a sample, so that it may obtain a highly
accurate measurement result that suppresses the impact of the aforementioned changes
in the interface.
A measuring apparatus according to a second embodiment of the present invention
will be described with reference to FIG.
3.
The measuring apparatus according to the second embodiment is a surface plasmon
sensor that utilize surface plasmon resonance as in the first embodiment, and FIG.
3 is a side view thereof. Elements identical to those of the apparatus shown in
FIG. 1 are given the same numerical symbols and will not be elaborated further here.
The measuring apparatus according to the second embodiment of the present invention
uses a measuring unit which is different in structure from that used in the measuring
apparatus shown in FIG.
1. More specifically, the measuring apparatus of
the second embodiment uses a measuring unit having a triangular prism
50
made of a dielectric material, which extends to the direction perpendicular to
the surface of FIG. 3, and a dielectric plate
55 connected to the upper
surface of the prism
50 through refractive index matching oil
52,
instead of using the measuring chip
10 used for the measuring apparatus
shown in FIG.
1. The prism
50 has an entrance surface
50a
and an outgoing surface
50b for a light beam L, while the dielectric
plate
55 has a metal film
56 and a transparent dielectric film
57
formed thereon in this order. The light beam L enters from the entrance surface
50a, and is totally reflected at the interface
55a between
the dielectric plate
55 and the metal film
56, and emerges from the
outgoing surface
50b. That is, the measuring unit of the second embodiment
uses a dielectric block that comprises a section
50 having the entrance
surface
50a and the outgoing surface
50b for the light
beam L, and a section
55 having the surface the metal film is formed thereon,
each formed as a separate body, and used as a measuring unit by connecting them
through the refractive index matching means (refractive index matching oil)
52.
Accordingly, a sample
58 placed on the transparent dielectric plate
55
is removable together with the dielectric plate
55 from the prism
50.
The measuring apparatus of the second embodiment enters the light beam into the
interface
55a between the dielectric plate
55 and the metal
film
56 by the light beam entering means
1 at various incident angles
within the angle range that satisfies the conditions of total reflection at the
interface and creates two dark lines, detects the positions of the two dark lines
by the light detecting means
17, and measures the distance (difference)
between the two dark lines to obtain the change in the dielectric constant of the
sample
58 placed on the transparent dielectric film
57, as in the
case of the measuring apparatus of the first embodiment. Accordingly, the measuring
apparatus according to the second embodiment may obtain a measurement result that
is not affected by the change in the inclination of the interface
55a
arising from, for example, resetting the dielectric block on the prism
50
through the matching oil
52 after a certain period of time from the time
when it was removed in order to efficiently carry out the measurement for a plurality
of samples.
In addition, if a sensing material is disposed on the transparent film
57,
as in the measuring apparatus of the first embodiment, the measuring apparatus
of the second embodiment may perform measurement of the combined state of the sensing
material and a particular substance contained in the sample solution injected thereon.
*
