JPH06109636A - Optical measuring device - Google Patents

Abstract

PURPOSE:To raise fluorescence intensity while suppressing the fluorescence intensity changes with lapse of time. CONSTITUTION:A laser light source 1 which emits measurement light, a condenser lens system 2 which condenses light flux before an incident surface 4a of a refractive coupling prism 4, a slab type optical waveguide channel 5 provided with the refractive coupling prism 4 at its end, a photo detector 7 which detects fluorescence, are provided. The measurement light condensed repeats total reflections while expanding in an optical waveguide channel, to increase detection area. The light flux is made to enter the slab type optical waveguide channel 5, while diffusing, so that the aging problem of fluorescence intensity, caused by destruction of an index fluorescence body 13a, is reduced.

Description

Detailed Description of the Invention

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to an optical measuring device, and more specifically, to a measurement existing in the vicinity of a reaction surface of an optical waveguide due to an evanescent wave component generated by guiding a measuring light through the optical waveguide. The present invention relates to an optical measuring device that measures optical characteristics of an object.

[0002]

2. Description of the Related Art Conventionally, an antigen, an antibody, or a hapten has been fixed in advance on the reaction surface of a slab type optical waveguide, and the evanescent wave component slightly exuding from the optical waveguide has been used to determine the amount of antigen-antibody reaction on the reaction surface of the optical waveguide. An immunoassay method for performing the measurement is known, and in order to embody this method, as shown in FIG. 7, a reaction tank 62 is integrally formed on one surface of a slab type optical waveguide 61, and a laser light source 60 emits the reaction light. The measuring light is introduced into the optical waveguide 61 through the dichroic mirror 63, the fluorescence emitted from the labeling fluorescent substance 68a is emitted through the optical waveguide 61, is reflected by the dichroic mirror 63, and is further passed through the optical filter 64 to be a photodetector. It has been proposed to make the light incident on 65 (Swiss patent application No. 2799/85).
-2 and JP-A-63-273042).

When the above configuration is adopted, the antibody 66 is previously fixed on the surface of the optical waveguide 61, and the antibody 6
6 receives the antigen 67 in the test liquid, and further the received antigen 67 receives the fluorescently labeled antibody 68 labeled with a fluorescent substance. That is, the amount of the fluorescent labeled antibody 68 received is determined based on the amount of the antigen 67 in the test liquid. Then, only the labeled fluorescent material 68a of the received fluorescent labeled antibody 68 is excited by the evanescent wave component generated by introducing the measurement light into the optical waveguide 61 and emits fluorescence, so that the intensity of the emitted fluorescence is measured. Antigen 67 in liquid
Will be proportional to the amount of. Further, this fluorescence is guided through the optical waveguide 61 as signal light.

Therefore, only the fluorescence guided through the optical waveguide 61 is reflected by the dichroic mirror 63 and is incident on the photodetector 65 through the optical filter 64 to measure the presence or absence of the immune reaction and the degree of the immune reaction. be able to.

[0005]

In the fluorescence immunoassay apparatus as shown in FIG. 7, a beam-shaped measuring light is made incident on the optical waveguide 61, and the measuring light is reflected by a minute area on the surface of the optical waveguide 61. While guiding inside the optical waveguide 61. Then, as the measurement light is guided, the evanescent wave is generated only in the vicinity of the region where the measurement light is reflected. Therefore,
Fluorescently labeled antibody 68 received on the surface of the optical waveguide 61
Since only some of the labeled fluorescent substances 68a are excited, the intensity of the fluorescence detected by the photodetector 65 cannot be sufficiently increased, and it is difficult to improve the sensitivity of the immunoassay. There was a problem.

As a simple method for increasing the intensity of fluorescence with respect to the above problems, (1) the intensity of measuring light is increased. (2) The incident angle when totally reflected by the surface of the optical waveguide is reduced. (3) Widen the measuring light. However, there are the following problems, and they are not effective countermeasures. (1) Problem of Method for Increasing Measurement Light Intensity When the measurement light intensity is increased, the fluorescence intensity is increased in proportion to the measurement light intensity. There is a limit to the increase in the intensity of the measurement light because it is destroyed and the intensity of fluorescence changes over time, and the accuracy of immunoassay decreases. (2) Problem of Method for Reducing Incident Angle when Reflected on Surface of Optical Waveguide When incident angle θ on the surface of optical waveguide 61 is decreased as shown in FIG. As shown in the figure, the number of reflections by the optical waveguide 61 having the same length increases as compared with the case where the incident angle θ is large, and therefore the excitation of the labeling fluorescent substance 68a of the fluorescent labeling antibody 68 received on the surface of the optical waveguide 61. The ratio of light emitted is increased and the amount of fluorescence is increased. However, the incident angle θ
Is smaller than the critical angle, the measurement light component 70 that is not totally reflected and is transmitted and is emitted into the test solution is generated, so that the measurement light to be guided is reduced, and as a result, the fluorescence amount is reduced. Further, due to the measurement light component 70 that passes through the test solution without being totally reflected, the fluorescent labeled antibody 68 suspended in a liquid other than the fluorescent labeled antibody 68 received on the surface of the optical waveguide 61. Since the labeled fluorescent substance 68a is also excited and fluorescence unrelated to the immune reaction is generated and the accuracy of the immunoassay is reduced, there is a limit to reducing the incident angle. (3) Problem of Method of Increasing Width of Measuring Light Increasing the width W of the measuring light increases the area where the measuring light is reflected on the surface of the optical waveguide 61 as shown in FIG. 9A. The ratio of excitation of the fluorescent labeled antibody 68 in the labeled fluorescent substance 68a received on the surface of the optical waveguide 61 increases, and the intensity of fluorescence increases. However, if the width W of the measurement light is made too wide, a part of the measurement light component 71 enters the test solution without being guided in the optical waveguide 61 as shown in FIG. Decrease the amount of fluorescence and
Fluorescently labeled antibody 6 that is not received on the surface of the optical waveguide 61
Since the labeled fluorescent substance 68a of No. 8 is excited, fluorescence unrelated to the antigen-antibody reaction is generated, and the accuracy of immunoassay is reduced. Therefore, there is a limit to the expansion of the width W of the measurement light.

[0007]

SUMMARY OF THE INVENTION The present invention has been made in view of the above problems, and provides an optical measuring device capable of improving the signal intensity of signal light showing the optical measurement result by the evanescent wave component of the measuring light. The purpose is to provide.

[0008]

The optical measuring device according to claim 1 for achieving the above object introduces the measuring light into the optical waveguide through a prism provided at a predetermined position of the optical waveguide, An optical measurement device that performs optical measurement near the surface of the optical waveguide by the evanescent wave component, emits signal light indicating the optical measurement result through the optical waveguide and prism, and guides it to the photodetector. It has an optical element that collects light at a position before the first reflecting surface of the waveguide.

An optical measuring device according to a second aspect is the optical measuring device according to the first aspect, which includes an optical system for converting the light emitted from the light source into a parallel light flux having a predetermined width. Claim 3
The optical measurement device according to claim 1, wherein the dichroic mirror for separating the measurement light and the signal light is provided between the light source and the optical waveguide, and the optical measurement device is provided between the dichroic mirror and the prism. An optical element is provided on the.

[0010]

In the optical measuring device according to the first aspect of the present invention, the measurement light is condensed by the optical element and then guided through the inside of the optical waveguide while diverging, whereby the area reflected on the surface of the optical waveguide gradually increases. Therefore, the surface of the optical waveguide on which optical measurement can be performed can be expanded, and the intensity of the signal light can be increased. Therefore, the signal light showing the optical measurement result can be increased to improve the measurement sensitivity. In addition, since the condensing position is located before entering the first reflecting surface of the optical waveguide, the intensity of the measurement light per unit area on the surface of the optical waveguide does not become high, and for example, a substance that emits signal light is It is also possible to suppress the problem that the intensity of the signal light is destroyed and changes over time.

According to the optical measuring device of the second aspect, when the light emitted from the light source is a beam light or a light which is not a parallel light flux (for example, divergent light), it is once converted into a parallel light flux having a predetermined width. The configuration in which the light is condensed by the optical element has an advantage that the design of the optical element and the adjustment of the condensing position of the optical element are simplified. According to the optical measuring device of claim 3, after the measuring light passes through the optical element through the dichroic mirror, the measuring light is made incident on the inside of the optical waveguide through the prism, and the evanescent wave component of the measuring light causes the light near the surface of the optical waveguide. Optical measurement is performed, and signal light showing the optical measurement result is emitted through the optical waveguide and the prism,
After passing through the optical element again, the light is guided to the photodetector via the dichroic mirror in an optical path different from the optical path of the measurement light. At this time, since the optical element is provided between the dichroic mirror and the prism, the surface of the optical waveguide capable of performing optical measurement can be widened, and the effect of increasing the intensity of signal light can be obtained, and the optical element can be obtained. When there is no signal light, the signal light in a direction that cannot be received by the photodetector can be collected by the optical element and made incident on the photodetector, so that the measurement sensitivity of the signal light can be further improved.

[0012]

Embodiments will be described in detail below with reference to the accompanying drawings showing embodiments. FIG. 1 is a schematic diagram showing the configuration of an immunoassay device as an embodiment of the optical measurement device of the present invention. This immunoassay device includes a laser light source 1 such as a semiconductor laser, a condenser lens system 2 for converging the beam light emitted from the laser light source 1 at a predetermined position after separating the measurement light and the signal light. A dichroic mirror 3 having the function of
A slab type optical waveguide 5 having a refraction coupling prism 4,
It has an optical filter 6 that cuts light in a predetermined wavelength region of the light reflected by the dichroic mirror 3 and a photodetector 7 that detects signal light with high sensitivity.

The condenser lens system 2 has a focal length set so that the measurement light emitted from the laser light source 1 is transmitted through the dichroic mirror 3 and condensed in front of the refraction-type coupling prism 4. The divergent measurement light is set to enter from the entrance surface 4a of the refractive coupling prism 4. A reaction tank 10 is formed on at least one surface of the slab type optical waveguide 5, and an antibody 11 is fixed on the surface facing the reaction tank 10.

The operation of the immunoassay device constructed as described above is as follows. First, the antibody 1 immobilized on the slab type optical waveguide 5
1 to receive the antigen 12 in the test liquid, and the received antigen 1
The point that the fluorescent labeled antibody 13 having the labeled fluorescent substance 13a on 2 is received is the same as the conventional immunoassay device. The beam light emitted from the laser light source 1 is made into a convergent light after the width of the light is widened by the condenser lens system 2, transmitted through the dichroic mirror 3 and provided in the slab type optical waveguide 5. The light is condensed in front of the entrance surface 4a of the prism 4. Then, in a state of diverging from the condensing position, the light enters the refractive coupling prism 4 from the incident surface 4a. The incident measurement light is guided while diverging from the refraction-type coupling prism 4, and reaches the slab-type optical waveguide 5.

Then, the measurement light guided into the slab type optical waveguide 5 is repeatedly spread and guided in the slab type optical waveguide 5 while being gradually spread, and is confined near the surface depending on the evanescent wave component. The labeled fluorescent substance 13a emits fluorescence. Then, the fluorescence is introduced into the slab type optical waveguide 5 and emitted from the refractive type coupling prism 4. The fluorescence emitted from the refraction-type coupling prism 4 is reflected by the dichroic mirror 3, passes through the optical filter 6 adjusted so that only the fluorescence is transmitted, removes noise light, and then enters the photodetector 7. .

Here, the condenser lens system 2 is condensed in front of the incident surface 4a of the refraction-type coupling prism 4 and is incident on the surface of the optical waveguide 5 in a state where light is diverged, whereby the surface of the optical waveguide 5 is made. The area reflected by the optical waveguide 5 gradually increases, and the ratio of the labeled fluorescent substance 13a excited in the labeled fluorescent substance 13a of the fluorescent labeled antibody 13 received on the surface of the optical waveguide 5 can be increased. The fluorescence as light can be increased. In addition, since the measurement light is already sufficiently diverged in the region S which is the first reflection surface of the optical waveguide 5, the intensity per unit area of the measurement light when reflected by the surface of the optical waveguide 5 is the labeled phosphor. It is not strong enough to destroy 13a.

FIG. 2 is a schematic diagram showing the construction of an immunoassay device showing a comparative example relating to the intensity of the measuring light per unit area. In the case of this comparative example, the condenser lens system 2
Although the measurement light is condensed by the same, the intensity of the measurement light per unit area increases because the condensed position is at the surface position of the optical waveguide 5 or a position close to the surface, and the labeled fluorescent substance It happens that 13a is destroyed. Therefore, there arises a problem that the fluorescence intensity changes with time and the accuracy of the immunoassay decreases.

FIG. 3 is a diagram showing data obtained by measuring changes with time in fluorescence intensity detected by the photodetector 7 in the configuration of the comparative example, in which the horizontal axis represents time and the vertical axis represents fluorescence intensity (arbitrary unit). ing. Further, FIG. 4 is a diagram showing data in the case where the condensing position of the measurement light is set in front of the entrance surface 4a of the refractive coupling prism 4 as shown in this embodiment. Comparing FIG. 3 and FIG. 4, it was found that the fluorescence intensity was the same in the initial state (time 0), but by adopting the configuration of this example,
It can be understood that the change in fluorescence intensity over time can be suppressed. This is because the ratio of the fluorescent light in the optical waveguide 5 is larger in the surface region of the optical waveguide 5 closer to the refractive coupling prism 4 than in the surface region of the optical waveguide 5 on the far side. It seems that this is because the measured light intensity per unit area in the surface region is not made stronger than the limit.

To summarize the above, in the configuration of the comparative example, the measurement light is condensed and is incident on the optical waveguide 5, so that the area reflected in the optical waveguide 5 can be gradually guided and guided in the optical waveguide. Since it is possible to increase the intensity of the fluorescence as the signal light, the intensity of the measurement light per unit area becomes too strong because the position of converging light is located near the surface of the optical waveguide 5, and There is a problem that it changes over time. On the other hand, in this embodiment, since the condensing position is in front of the entrance surface 4a of the refraction-type coupling prism 4, the labeled fluorescent substance is retained while maintaining the advantage of increasing the intensity of fluorescence due to the increase of the area. It is possible to reduce the problem of time-dependent change in fluorescence intensity due to destruction of 13a.

[0020]

[Embodiment 2] FIG. 5 is a schematic diagram showing the structure of an immunoassay device as another embodiment of the optical measurement device of the present invention.
This embodiment is different from the above-mentioned embodiment only in that the beam light emitted from the laser light source 1 is expanded by the beam expander 20 and then condensed by the condenser lens 2a at a predetermined position.

The configuration of this embodiment also has the same effect as that of the above-described embodiment, and the light beam is once spread to form a parallel light beam having a predetermined width, and then the light beam is condensed by the condenser lens 2a. There is an advantage that the design of the condenser lens 2a itself is simplified and the adjustment work of the condensing position of the measurement light in each optical measuring device can be easily performed.

[0022]

[Embodiment 3] FIG. 6 is a schematic diagram showing the structure of an immunoassay device as another embodiment of the optical measurement device of the present invention.
This embodiment is different from the second embodiment in that a condenser lens 2b is provided between the dichroic mirror 3 and the refraction-type coupling prism 4 so that the beam expander 20 collimates a light beam having a predetermined width. The only difference is that after passing through the mirror 3, the light is condensed by the condenser lens 2b in front of the entrance surface 4b of the refractive coupling prism 4. With this structure, the same effect as that of the first embodiment can be obtained, and the fluorescence as the signal light emitted from the refractive coupling prism 4 can be collected over a wide range by the condenser lens 2b. ,
There is an advantage that the condensing property of fluorescence can be improved and the measurement accuracy can be improved.

However, in order to prevent the measurement light from being reflected on the condenser lens surface 2b1 on the dichroic mirror 3 side, it is necessary to provide an antireflection film on the condenser lens surface 2b1 to reduce the signal light S /
It is preferable for improving the N ratio. The present invention is not limited to the above embodiments, and various design changes can be made without departing from the scope of the present invention.

In the present invention, the effect of the present invention can be obtained if the measuring light is made to enter the region S which is the first reflecting surface of the slab type optical waveguide 5 in a state of being diverged to some extent. Therefore, the condensing position is not limited to the front of the incident surface 4b of the refractive coupling prism 4. For example, the condensing position is set according to the shape of the refraction coupling prism 4 and the distance between the refraction coupling prism 4 and the optical waveguide 5, and is set so as not to cause unnecessary reflection in the refraction coupling prism 4. By
It is also possible to set the condensing position of the condensing lens system 2 or the condensing lenses 2a and 2b in the refractive coupling prism 4. In this case as well, the light diverging from the condensing position in the refractive coupling prism 4 is incident on the optical waveguide 5.

In the above embodiment, the case where the laser light source 1 such as a semiconductor laser is used as the light source has been described as an example. However, a light source such as a halogen lamp is monochromatic by a filter and a light beam is narrowed by a slit. It is obvious that the present invention can be similarly applied to an optical measuring device that employs an optical system that makes the light incident on the optical waveguide 5.

Furthermore, in the above-described embodiment, the optical axis on which the measurement light is incident and the optical axis on which the signal light is measured are coincident with each other, and the configuration in which the measurement light and the signal light are separated by the dichroic mirror 3 has been described. The present invention can be similarly applied to an optical measuring device that does not use the dichroic mirror 3 and has a configuration in which the optical axis on which the measurement light is incident does not match the optical axis on which the signal light is measured.

[0027]

As described above, according to the first aspect of the present invention, the change over time of the signal light intensity is achieved by the simple structure in which the optical element for condensing the measurement light at the position before the first reflecting surface of the optical waveguide is provided. It has a peculiar effect that the signal light intensity is improved by increasing the area reflected in the optical waveguide while reducing

In addition to the effect of claim 1, the invention of claim 2 has a unique effect that the design of the optical element itself and the adjustment of the converging position of the optical element in each optical measuring device are simplified. According to the invention of claim 3, in addition to the effect of claim 1, the signal light emitted from the prism is collected by the optical element and is incident on the photodetector, so that the signal light intensity can be further improved. Produce the effect of.

[Brief description of drawings]

FIG. 1 is a schematic diagram showing the configuration of an immunoassay device as an embodiment of the optical measurement device of the present invention.

FIG. 2 is a schematic diagram showing a configuration of an immunoassay device of a comparative example.

FIG. 3 is a diagram showing data obtained by measuring changes in fluorescence intensity over time in a comparative example.

FIG. 4 is a diagram showing data obtained by measuring changes in fluorescence intensity over time in Examples of the present invention.

FIG. 5 is a schematic diagram showing the configuration of an immunoassay device as another embodiment of the optical measurement device of the present invention.

FIG. 6 is a schematic diagram showing the configuration of an immunoassay device as another embodiment of the optical measurement device of the present invention.

FIG. 7 is a diagram showing an example of a configuration of a conventional immunoassay device.

FIG. 8 is a diagram for explaining a problem of a method of reducing an incident angle when reflected on the surface of an optical waveguide.

FIG. 9 is a diagram for explaining a problem of the method of widening the width of the measurement light.

[Explanation of symbols]

1 laser light source 2 condenser lens system 2a, 2b
Condensing lens 3 Dichroic mirror 4 Refractive coupling prism 5 Slab type optical waveguide 7 Photodetector 20 Beam expander

Claims (3)

[Claims] 1. The measuring light is introduced into the optical waveguide (5) through a prism (4) provided at a predetermined position of the optical waveguide (5), and the evanescent wave component of the measuring light causes the vicinity of the surface of the optical waveguide (5). Of the optical waveguide (5) and the prism (4).
An optical measuring device that emits light through the optical detector (7) and guides it to the photodetector (7), and collects the measuring light at a position before the first reflecting surface of the optical waveguide (5) (2) (2a) ( 2b)
An optical measuring device comprising: 2. The optical measuring device according to claim 1, further comprising an optical system (20) for collimating the light emitted from the light source (1) into a parallel light flux having a predetermined width. 3. A dichroic mirror (3) for separating measurement light and signal light between a light source (1) and an optical waveguide (5).
And an optical element (2b) between the dichroic mirror (3) and the prism (4).
The optical measuring device according to.