CN101451995A - Optical-waveguide sensor chip, method of manufacturing the same, method of measuring substance, substance-measuring kit and optical-waveguide sensor - Google Patents

Abstract

The invention aims to provide an optical-waveguide sensor for measuring an object substance of tested samples quantitatively with less tested sample amount and in shorter period. It is realized by the following method: an optical-waveguide sensor chip includes an optical waveguide having a first substance immobilized on the surface thereof, the first substance being specifically reactive with an analyte substance, and fine particles dispersed on the optical waveguide and having a second substance immobilized on the surface thereof, the second substance being specifically reactive with the analyte substance.

Description

The assay method of optical-waveguide sensor chip and manufacture method thereof, material, substance-measuring kit and optical-waveguide sensor

Technical field

The present invention relates to the manufacture method of optical-waveguide sensor chip, optical-waveguide sensor chip, assay method, substance-measuring kit and the optical-waveguide sensor of material.

Background technology

The existing immunoassays that utilize antigen-antibody reaction usually will be corresponding with protein in the determined sample etc. an antibody immobilization at poroid substrate surface.In the hole, drip determined sample solution, secondary antibodies liquid, the chromogenic reagent of ormal weight respectively successively.The cleansing solution washing that each dropping of each solution all will be stipulated respectively.Such immunoassays all need mensuration person to carry out with the process of the complexity of weighing one after another, interpolation and discharge and so on.In such immunoassays, the capacity of determined sample needs about 5 μ L～25 μ L at least.

On the other hand, the amount of the determined sample that the minimum of the applicant in the patent documentation of having applied for 1 is essential is on the basis of 1 μ L, even it is incorrect to disclose the capacity of determined sample, also can carry out the method for measurement of concentration of concentration determination of the determination object material of determined sample, and sensor chip.

Patent documentation 1:WO2005/022155

But, in existing immunoassay system, the reaction of antibody and the reaction of determined sample, determined sample and secondary antibodies all needs about one hour respectively, in addition, also need process, therefore exist from determined sample collecting and need the problem of several hrs to obtaining measurement result through previously described complexity.In addition, because the amount of the determined sample that needs is many, therefore in the blood test of using toys such as rat for example, the inspection item that carries out several kinds for 1 time just needs to sacrifice 1 sample.Consequently be difficult to check same sample over time.

Summary of the invention

The present invention has carried out further improvement to the invention of above-mentioned patent documentation 1, but provide with still less determined sample size, in the shorter time assay method of manufacture method, the material of optical-waveguide sensor chip, the optical-waveguide sensor chip of the determination object material of the determined sample of quantitative measurement and substance-measuring kit, optical-waveguide sensor.

According to the 1st mode of the present invention, a kind of optical-waveguide sensor chip is provided, it is characterized in that it has:

Immobilization has the optical waveguide with the 1st material of determination object material specific reaction from the teeth outwards; With,

Be dispersed on the above-mentioned optical waveguide and immobilization has particulate with the 2nd material of said determination object material specific reaction.

According to the 2nd mode of the present invention, a kind of optical-waveguide sensor chip is provided, it is characterized in that it has:

Immobilization has the optical waveguide with the 1st material of determination object material specific reaction from the teeth outwards;

The back up pad that disposes in opposite directions with above-mentioned optical waveguide; With,

What be scattered in above-mentioned back up pad has particulate with the 2nd material of said determination object material specific reaction with surface and the immobilization in opposite directions of above-mentioned optical waveguide.

According to the 3rd mode of the present invention, a kind of manufacture method of optical-waveguide sensor chip is provided, it is characterized in that, comprise the steps:

Will be with determination object material specific reaction the 1st substance fixed in light guide surface;

To contain immobilization has with the slurry of the particulate of the 2nd material of said determination object material specific reaction and coats on the above-mentioned optical waveguide; With,

Dry after above-mentioned coating, with above-mentioned microparticulate on above-mentioned optical waveguide.

According to the 4th mode of the present invention, a kind of manufacture method of optical-waveguide sensor chip is provided, it is characterized in that, comprise the steps:

Will be with determination object material specific reaction the 1st substance fixed in light guide surface;

To contain immobilization has with the slurry of the particulate of the 2nd material of said determination object material specific reaction and coats the back up pad surface;

Dry after above-mentioned coating, with above-mentioned microparticulate in above-mentioned back up pad; With,

With above-mentioned back up pad according to its microparticulate face and above-mentioned optical waveguide mode are in opposite directions disposed spaced apartly.

According to the 5th mode of the present invention, a kind of assay method of material is provided, it is characterized in that, comprise the steps:

Prepare optical-waveguide sensor chip, this optical-waveguide sensor chip has: immobilization has with the optical waveguide of the 1st material of determination object material specific reaction and is dispersed on the above-mentioned optical waveguide and immobilization has particulate with the 2nd material of said determination object material specific reaction from the teeth outwards;

Light guide surface to above-mentioned sensor chip drips determined sample solution, make between the 1st material of light guide surface and the determination object material in the determined sample solution specific reaction takes place, make said determination object material simultaneously and be dispersed between the 2nd material of the particulate on the above-mentioned optical waveguide specific reaction takes place; With

Detect to be situated between and to be immobilized onto the optical change that particulate produced of above-mentioned light guide surface by the 1st material and determination object material.

According to the 6th mode of the present invention, a kind of assay method of material is provided, it is characterized in that, comprise the steps:

Prepare optical-waveguide sensor chip, this optical-waveguide sensor chip has immobilization from the teeth outwards optical waveguide with the 1st material of determination object material specific reaction is arranged;

Light guide surface to above-mentioned sensor chip drips determined sample solution, makes between the 1st material of light guide surface and the determination object material in the determined sample solution specific reaction takes place;

Wash above-mentioned light guide surface;

Dripping immobilization to above-mentioned light guide surface has dispersion liquid with the particulate of the 2nd material of the determination object material specific reaction of determined sample, makes between the 2nd material of the determination object material of determined sample solution and particulate specific reaction takes place; With,

Detect to be situated between and to be immobilized onto the optical change that particulate produced of light guide surface by the 1st material and determination object material.

According to the 7th mode of the present invention, a kind of assay method of material is provided, it is characterized in that, comprise the steps:

Prepare optical-waveguide sensor chip, this optical-waveguide sensor chip has immobilization from the teeth outwards optical waveguide with the 1st material of determination object material specific reaction is arranged;

Being pre-mixed determined sample solution and immobilization has particulate with the 2nd material of the determination object material specific reaction of determined sample, makes the 2nd material of particulate and the determination object material generation specific reaction of determined sample solution;

Light guide surface to above-mentioned sensor chip drips above-mentioned mixed liquor, makes the 1st material of above-mentioned light guide surface and carries out specific reaction with the determination object material of the determined sample solution of the 2nd substance reaction of particulate; With,

Detect to be situated between and to be immobilized onto the optical change that particulate produced of above-mentioned light guide surface by the 1st material and determination object material.

According to the 8th mode of the present invention, a kind of assay method of material is provided, it is characterized in that, comprise the steps:

Prepare optical-waveguide sensor chip, this optical-waveguide sensor chip has immobilization from the teeth outwards optical waveguide with the 1st material of determination object material specific reaction is arranged;

Light guide surface to above-mentioned sensor chip drips determined sample solution, makes between the 1st material of light guide surface and the determination object material in the determined sample solution specific reaction takes place;

Dripping immobilization to above-mentioned light guide surface has dispersion liquid with the particulate of the 2nd material of the determination object material specific reaction of determined sample, makes between the 2nd material of said determination object material and particulate specific reaction takes place; With,

Detect to be situated between and to be immobilized onto the optical change that particulate produced of above-mentioned light guide surface by the 1st material and determination object material.

According to the 9th mode of the present invention, a kind of assay method of material is provided, it is characterized in that, comprise the steps:

Prepare optical-waveguide sensor chip, this optical-waveguide sensor chip has immobilization from the teeth outwards optical waveguide with the 1st material of determination object material specific reaction is arranged;

Drip immobilization to the light guide surface of above-mentioned sensor chip dispersion liquid with the particulate of the 2nd material of the determination object material specific reaction of determined sample is arranged;

There is the above-mentioned light guide surface of above-mentioned dispersion liquid to drip determined sample solution to dropping, make between the 1st material of light guide surface and the determination object material in the determined sample solution specific reaction takes place, make simultaneously between the 2nd material of the particulate in said determination object material and the above-mentioned dispersion liquid specific reaction takes place; With,

Detect to be situated between and to be immobilized onto the optical change that particulate produced of described light guide surface by the 1st material and determination object material.

According to the 10th mode of the present invention, a kind of assay method of material is provided, it is characterized in that, comprise the steps:

Prepare optical-waveguide sensor chip, this optical-waveguide sensor chip has: immobilization has the optical waveguide with the 1st material of determination object material specific reaction from the teeth outwards, the back up pad that disposes in opposite directions with above-mentioned optical waveguide and be scattered in above-mentioned back up pad with surface and the immobilization in opposite directions of above-mentioned optical waveguide particulate with the 2nd material of said determination object material specific reaction is arranged;

Between the optical waveguide of above-mentioned sensor chip and above-mentioned back up pad, inject determined sample solution, make between the 1st material of light guide surface and the determination object material in the determined sample solution specific reaction takes place, make said determination object material simultaneously and be scattered between the 2nd material of particulate of above-mentioned back up pad specific reaction takes place; With

Detect to be situated between and to be immobilized onto the optical change that particulate produced of described light guide surface by the 1st material and determination object material.

According to the 11st mode of the present invention, a kind of substance-measuring kit is provided, it is characterized in that its combination has optical-waveguide sensor chip and package body,

Described optical-waveguide sensor chip has optical waveguide and lid, described optical waveguide immobilization from the teeth outwards has the 1st material with the determination object material specific reaction of determined sample, described lid is disposed at described light guide surface, and described light guide surface between have and be used to form the recess of measuring the district, and opening has entrance hole and the exhaust opening that is communicated with this mensuration district;

Described package body is taken in immobilization dispersion liquid with the particulate of the 2nd material of the determination object material specific reaction of determined sample.

According to the 12nd mode of the present invention, a kind of assay method of material is provided, it is characterized in that, wherein use above-mentioned substance to measure and use kit, and comprise the steps:

Determined sample solution is added drop-wise to the light guide surface of measuring in the district via the entrance hole of optical-waveguide sensor chip lid, makes the 1st material that is immobilized onto light guide surface and the determination object material generation specific reaction of determined sample solution;

The dispersion liquid of particulate is imported to the light guide surface of measuring in the district via the entrance hole of above-mentioned lid, simultaneously, during determined sample solution is discharged via exhaust opening, make between the 2nd material of the determination object material of the determined sample solution that specific reaction takes place and particulate specific reaction takes place; With,

Detect to be situated between and to be immobilized onto the optical change that particulate produced of light guide surface by the 1st material and determination object material.

According to the 13rd mode of the present invention, a kind of optical-waveguide sensor is provided, it is characterized in that it has:

Optical-waveguide sensor chip, it has: immobilization has with the optical waveguide of the 1st material of determination object material specific reaction and is dispersed on the described optical waveguide and immobilization has particulate with the 2nd material of described determination object material specific reaction from the teeth outwards;

Make light incide the light source of above-mentioned optical waveguide; With,

Reception is from the photo detector of the light of above-mentioned optical waveguide ejaculation.

According to the 14th mode of the present invention, a kind of optical-waveguide sensor is provided, it is characterized in that it has:

Optical-waveguide sensor chip, it has: immobilization has the optical waveguide with the 1st material of determination object material specific reaction from the teeth outwards, the back up pad that disposes in opposite directions with described optical waveguide, and be scattered in described back up pad with surface and the immobilization in opposite directions of described optical waveguide particulate with the 2nd material of described determination object material specific reaction is arranged;

Make light incide the light source of above-mentioned optical waveguide; With,

Reception is from the photo detector of the light of above-mentioned optical waveguide ejaculation.

According to the present invention, can provide can be still less determined sample size, in the shorter time, the optical-waveguide sensor of the determination object material of the determined sample of quantitative measurement, the manufacture method and the optical-waveguide sensor of optical-waveguide sensor chip.

In addition, according to the present invention, can provide can be still less determined sample size, in the shorter time, the assay method of the material of the determination object material of quantitative determined sample.

And then, according to the present invention, can provide can be still less determined sample size, in the shorter time, the substance-measuring kit of the determination object material of the determined sample of quantitative measurement.

Description of drawings

Fig. 1 is the sectional view of optical-waveguide sensor that expression has the optical-waveguide sensor chip of the 1st embodiment.

Fig. 2 is the skeleton diagram of mensuration operation of determination object material of the determined sample of expression the 1st embodiment.

Fig. 3 represents to have the sectional view of optical-waveguide sensor of the optical-waveguide sensor chip of the 2nd embodiment.

Fig. 4 represents the skeleton diagram of mensuration operation of the material of the 2nd embodiment.

Fig. 5 represents the sectional view of the optical-waveguide sensor chip that uses in the assay method of material of the 3rd embodiment.

Fig. 6 represents the skeleton diagram of mensuration operation of the material of the 3rd embodiment.

Fig. 7 represents the skeleton diagram of mensuration operation of the material of the 4th embodiment.

Fig. 8 represents the sectional view of optical-waveguide sensor chip of the substance-measuring kit of the 5th embodiment.

Fig. 9 represents the skeleton diagram of mensuration operation of the material of the 5th embodiment.

The characteristic pattern that changes of laser light intensity as time passes during the concentration of insulin that Figure 10 is illustrated in embodiment 1 is measured.

The characteristic pattern that changes of laser light intensity as time passes during the concentration of insulin that Figure 11 is illustrated in embodiment 2 is measured.

The characteristic pattern that changes of laser light intensity as time passes during the concentration of insulin that Figure 12 is illustrated in embodiment 3 is measured.

The characteristic pattern that changes of laser light intensity as time passes during the concentration of insulin that Figure 13 is illustrated in embodiment 4 is measured.

Symbol description

1,31 glass substrates

2a, 2b, 32a, 32b grating

3,33 planar optical waveguides

5 reacting holes

11 the 1st materials

12 the 2nd materials

13 particulates

15 determination object materials

34 lids

35 measure the district

37 entrance holes

38 exhaust openings

Embodiment

Below, as embodiment of the present invention, describe optical-waveguide sensor, the manufacture method of optical-waveguide sensor chip, assay method, substance-measuring kit and the optical-waveguide sensor of material in detail.

(the 1st embodiment)

The optical-waveguide sensor chip of the 1st embodiment has: immobilization has with the optical waveguide of the 1st material of determination object material specific reaction and is scattered in this optical waveguide and immobilization has particulate with the 2nd material of said determination object material specific reaction from the teeth outwards.

Here, the determination object material for example can list protein contained in blood, serum, blood plasma, Biosample, the food etc., polypeptide, gene etc.Particularly, can list insulin, casein, beta lactoglobulin, ovalbumin, calcitonin, the C-polypeptide, leptin (leptin), beta-2-microglobulin, RBP ELISA, α-1-microglobulin, alfafoetoprotein, carcinomebryonic antigen, troponin-i, glucagon kind polypeptide, the Insulin-Like polypeptide, TGF, fibroblast growth factor, PDGF, epidermal growth factor, cortisol, trilute, haptens hormones such as thyroxine, digoxin, medicines such as theophylline, bacterium, infectious substances such as virus, hepatitis antibody, IgE, and the main protein complex that comprises buckwheat, the soluble protein of the Arah2 of peanut etc. etc., but be not limited to these.In addition, the determination object material in the 2nd embodiment to the 5 embodiments also uses same material below.

Optical waveguide for example can be used planar optical waveguide.This planar optical waveguide for example can be formed by the heat-curing resin or the alkali-free glass of phenolics, epoxy resin etc.At length, material used herein is meant the material of the optical transmission with regulation, especially preferably is the epoxy resin etc. of primary structure with the polystyrene.With the immobilization of the 1st material of the determination object material specific reaction of determined sample, for example utilizing silane coupling agent etc. to carry out carrying out immobilization by the hydrophobic interaction of above-mentioned substance on the surface that hydrophobization handles to planar optical waveguide.The 1st material is under the situation of antigen at the determination object material of determined sample for example, can use antibody.

Here " particulate is scattered in optical waveguide " is meant that particulate is scattered in light guide surface directly or indirectly.The mode of " particulate is scattered in light guide surface indirectly " for example can list particulate Jie is scattered in light guide surface by confining bed mode.Confining bed for example contains the water-soluble substances of polyvinyl alcohol (PVA), bovine serum albumin(BSA) (BSA), polyglycol, phospholipid polyalcohol, gelatin, carbohydrate (for example sucrose, trehalose) etc.Confining bed can further contain protein inhibitor.

Particulate for example can use the inorganic oxide particle of the metallic colloid of the resin beads of LATEX BEAD (trade name) of POLYSTYRENE system etc. or collaurum etc. or Titanium particles etc.Particulate also can use the nonmetal particle of polysaccharide, silicon dioxide granule, carbon particle of protein, agarose of albumin etc. etc. etc.Preferred especially LATEX BEAD, metallic colloid.Among the LATEX BEAD, when the light that conducts in the aftermentioned optical waveguide is red laser, preferred blue LATEX BEAD.

Particulate preferably has the diameter of 50nm～10 μ m.

The 2nd material is under the situation of antigen at the determination object material of determined sample for example, can use antibody.

Below, the manufacture method of the optical-waveguide sensor chip of the 1st embodiment is described.

At first, will be with determination object material specific reaction the 1st substance fixed on light guide surface.Then, by for example physisorption or be situated between by the chemical bond of carboxyl or amino etc. will be with the determination object material specific reaction of determined sample the 2nd substance fixed to particulate.Then, the microparticulate that immobilization is had the 2nd material is in containing the physiological saline of water-soluble substances, with the preparation slurry.This slurry is applied to after the optical waveguide dry, with above-mentioned microparticulate on above-mentioned optical waveguide, thereby make optical-waveguide sensor chip.

In such manufacture method, water-soluble substances for example can use polyvinyl alcohol (PVA), bovine serum albumin(BSA) (BSA), polyglycol, phospholipid polyalcohol, gelatin, carbohydrate (for example sucrose, trehalose).In addition, in order to improve the dispersiveness of particulate, dry preferably freeze drying.

Specifically describe the optical-waveguide sensor chip of the 1st embodiment with reference to Fig. 1.Fig. 1 is the sectional view of the optical-waveguide sensor chip of expression the 1st embodiment.

Both ends at the principal plane of glass substrate 1 are provided with incident sidelight grid 2a and emitting side grating 2b.These gratings 2a, 2b for example can be by titanium dioxide (TiO ₂), tin oxide (SnO ₂), formation such as zinc paste, lithium niobate, gallium arsenide (GaAs), indium tin oxide (ITO), polyimide.For example the planar optical waveguide 3 that is made of heat-curing resin forms on substrate 1 principal plane that comprises grating 2a, 2b.Low refractive index resin film 4 covers on the planar optical waveguide 3.Low refractive index resin for example can use poly-(the perfluorobuttene base vinyl ether) etc. of the CYTOP (registered trademark) of commercially available Asahi Glass Co., Ltd's system.In 4 upper sheds of low refractive index resin film,, for example form rectangular reacting hole 5 so that the part of the planar optical waveguide 3 between grating 2a, 2b is exposed.The cell wall 6 of frame shape is formed on the low refractive index resin film 4 mode of the reacting hole 5 that planar optical waveguide 3 exposes according to encirclement.

With the 1st material 11 of the determination object material specific reaction of determined sample, by the hydrophobization that for example utilizes silane coupling agent handle and immobilization to from reacting hole (measure and distinguish) 5 planar optical waveguides of exposing, 3 surfaces.Immobilization has the particulate 13 with the 2nd material 12 of the determination object material specific reaction of determined sample to be dispersed to immobilization and has on planar optical waveguide 3 surfaces of above-mentioned substance 11.The dispersion of this particulate 13 for example is applied on the planar optical waveguide 3 by the slurry that will contain particulate and water-soluble substances and freeze drying forms.

The optical-waveguide sensor of the 1st embodiment has: be used to make light to incide the light source (for example red laser diode) 21 of planar optical waveguide 3 and receive the photo detector (for example photodiode) 22 of the light that penetrates from emitting side grating 2b from the light incident side grating 2a of above-mentioned optical-waveguide sensor chip.

Below, explanation uses above-mentioned optical waveguide sensor to carry out the method for substance-measuring with reference to Fig. 2 (A)～(C).

At first, the optical-waveguide sensor chip shown in the set-up dirgram 2 (A).This sensor chip comprises the substrate 1 with grating 2a, 2b.Planar optical waveguide 3 forms on substrate 1 principal plane that comprises grating 2a, 2b.Low refractive index resin 4 covers on the planar optical waveguide 3, and according to the mode opening that a part that makes the planar optical waveguide 3 between grating 2a, 2b is exposed, for example forms rectangular reacting hole 5.Be immobilized from planar optical waveguide 3 surfaces that reacting hole 5 exposes with the 1st material (for example the 1st antibody) 11 of determination object material (for example antigen) specific reaction of determined sample.Immobilization has a plurality of particulates 13 with the 2nd material (for example the 2nd antibody) 12 of the determination object material specific reaction of determined sample to be dispersed on the planar optical waveguide 3.

Then, planar optical waveguide 3 surfaces of the discrete areas of the above-mentioned particulate 13 in comprising reacting hole 5 drip determined sample solution.At this moment, if do not exist in the determined sample solution that drips with the 1st antibody 11 on planar optical waveguide 3 surfaces with the 2nd antibody 12 of particulate 13 antigen of specific reactions take place, then shown in Fig. 2 (B), the 2nd antibody 12 of particulate 13 does not combine with the 1st antibody 11 on planar optical waveguide 3 surfaces, but is dispersed in the determined sample solution 14.Under this state, make red laser incide planar optical waveguide 3 from light incident side grating 2a from red laser diode 21, near surface (exposing surfaces at reacting hole 5 places), send the light that dies in wink (evanescent light) even in this planar optical waveguide 3, conduct, because the particulate 13 in the determined sample solution 14 in the reacting hole 5 disperses, particulate 13 also exists in the light zone that dies in wink hardly.That is to say that particulate 13 is irrelevant substantially with the absorption or the scattering of the light that dies in wink, the decay of the light intensity that therefore also takes place hardly to die in wink.Consequently, the red laser that penetrates from emitting side grating 2b receives the light time at photodiode 22, and its laser light intensity does not almost change.

On the other hand, if have antigen in the determined sample solution 14 that drips, then shown in Fig. 2 (C), antigen-antibody reactions take place with the 1st antibody 11 on planar optical waveguide 3 surfaces and combine in antigen 15, and then the 2nd antibody 12 of particulate 13 is with antigen 15 generation antigen-antibody reactions and combine.That is, between the 2nd antibody 12 of the 1st antibody 11 on planar optical waveguide 3 surfaces and particulate 13, be situated between and antigen-antibody reactions take place, so particulate 13 is immobilized on planar optical waveguide 3 surfaces by antigen 15.

After above-mentioned determined sample solution just drips, red laser from red laser diode 21 is incided on the planar optical waveguide 3 from light incident side grating 2a, near surface (exposing surfaces at reacting hole 5 places), send the light that dies in wink if in this planar optical waveguide 3, conduct, then be immobilized on planar optical waveguide 3 surfaces, so particulate 13 exists at the light region memory that dies in wink owing to particulate 13.That is to say that particulate 13 is relevant with the absorption or the scattering of the light that dies in wink, the decay of the light intensity that therefore takes place to die in wink.Consequently, the red laser that penetrates from emitting side grating 2b receives the light time at photodiode 22, and its laser light intensity is because the influence of immobilized particulate 13 and along with the time reduces.

The reduced rate of the laser light intensity that receives at photodiode 22 places and the amount of the particulate 13 that is immobilized onto planar optical waveguide 3 surfaces, promptly the antigen concentration in the relevant determined sample solution 14 is proportional with antigen-antibody reaction.Therefore, be produced in the known determined sample solution of antigen concentration the reduction curve of laser light intensity as time passes, obtain this curve and locate the reduced rate of laser light intensity at the appointed time, make the typical curve that concerns between the reduced rate of expression antigen concentration and laser light intensity in advance.From obtaining the reduced rate of laser light intensity at the appointed time with the time of said method mensuration and the reduction curve of laser light intensity, with the reduced rate of this laser light intensity and the contrast of above-mentioned typical curve, thereby can measure antigen concentration in the determined sample solution.

In addition, in above-mentioned concentration determination by making particulate and water-soluble substances be scattered in planar optical waveguide jointly, thereby improve the dispersiveness of particulate.In addition, when planar optical waveguide dripped determined sample solution, the water-soluble substances dissolving with particulate coexistence made particulate to move, thereby the reaction of the 2nd material of determination object material in the determined sample solution and particulate is carried out smoothly.

More than, according to the 1st embodiment, can provide minimum essential determined sample size few (for example 10 μ L are following), and can carry out quantitative optical-waveguide sensor chip, its manufacture method and optical-waveguide sensor to the concentration of the determination object material of determined sample by determined sample drop being added to the single job of measuring the district.

In addition, according to the 1st embodiment, can provide minimum essential determined sample size few (for example 10 μ L are following), and determined sample drop is added to the assay method that the single job of measuring the district can be carried out quantitative material to the concentration of the determination object material of determined sample.

(the 2nd embodiment)

The optical-waveguide sensor chip of the 2nd embodiment has: immobilization has the optical waveguide with the 1st material of determination object material specific reaction from the teeth outwards, the back up pad that disposes in opposite directions with this optical waveguide, and be scattered in this back up pad with surface and the immobilization in opposite directions of above-mentioned optical waveguide particulate with the 2nd material of said determination object material specific reaction is arranged.

Optical waveguide for example can be used planar optical waveguide.What illustrate in this planar optical waveguide and the 1st embodiment is same, can be formed by heat-curing resin or alkali-free glass.

Will with the 1st material of the determination object material specific reaction of determined sample with the same method immobilization that illustrates in the 1st embodiment on planar optical waveguide.The 1st material is under the situation of antigen at the determination object material of determined sample for example, can use antibody.

What illustrate in particulate and the 1st embodiment is same, for example can use the inorganic oxide particle of the metallic colloid of the resin beads of LATEX BEAD etc. or collaurum etc. or Titanium particles etc. etc.Particulate also can use the nonmetal particle of polysaccharide, silicon dioxide granule, carbon particle of protein, agarose of albumin etc. etc. etc.Preferred especially LATEX BEAD, metallic colloid.Particulate preferably has the diameter of 50nm～10 μ m.

The 2nd material is under the situation of antigen at the determination object material of determined sample for example, can use antibody.

Below, the manufacture method of the optical-waveguide sensor chip of the 2nd embodiment is described.

At first, will be with determination object material specific reaction the 1st substance fixed on light guide surface.Then, by for example physisorption or be situated between by the chemical bond of carboxyl or amino etc. will be with the determination object material specific reaction of determined sample the 2nd substance fixed to particulate.Then, the microparticulate that immobilization is had the 2nd material is in the physiological saline that contains water-soluble substances, with the preparation slurry.This slurry is applied to behind the back up pad surface dry, with above-mentioned microparticulate on back up pad.Thereafter, by with back up pad according to its microparticulate face and optical waveguide mode are in opposite directions disposed spaced apartly, thereby make optical-waveguide sensor chip.

In such manufacture method, water-soluble substances for example can use polyvinyl alcohol (PVA), bovine serum albumin(BSA) (BSA), polyglycol, phospholipid polyalcohol, gelatin, carbohydrate (for example sucrose, trehalose).In addition, in order to improve the dispersiveness of particulate, dry preferably freeze drying.

Specifically describe the optical-waveguide sensor of the 2nd embodiment with reference to Fig. 3.Fig. 3 is the sectional view of the optical-waveguide sensor chip of expression the 2nd embodiment.

Both ends at the principal plane of glass substrate 1 are provided with light incident side grating 2a and the emitting side grating 2b that for example is made of titanium dioxide.For example the planar optical waveguide 3 that is made of heat-curing resin forms on substrate 1 principal plane that comprises grating 2a, 2b.Low refractive index resin 4 covers on the planar optical waveguide 3.In 4 upper sheds of low refractive index resin film,, for example form rectangular reacting hole 5 so that the part of the planar optical waveguide 3 between grating 2a, 2b is exposed.For example the back up pad 7 that is made of synthetic resin forms according to the mode that covers reacting hole 5 on low refractive index resin film 4.The dropping of determined sample solution penetratingly is provided with from back up pad 7 surfaces towards reacting hole 5 with hole (not shown).

With the 1st material 11 of the determination object material specific reaction of determined sample, handle to be immobilized in from planar optical waveguide 3 surfaces that reacting hole 5 (measuring the district) exposes by the hydrophobization that for example utilizes silane coupling agent.Immobilization have the particulate 13 with the 2nd material 12 of the determination object material specific reaction of determined sample be dispersed to reacting hole 5 expose and with planar optical waveguide 3 back up pad 7 surfaces (lower surface) in opposite directions on.The dispersion of this particulate 13 for example coats above-mentioned back up pad surface by the slurry that will contain particulate and water-soluble substances and freeze drying forms.

The optical-waveguide sensor of the 2nd embodiment has: be used to make light to incide the light source (for example red laser diode) 21 of planar optical waveguide 3 and receive the photo detector (for example photodiode) 22 of the light that penetrates from emitting side grating 2b from the light incident side grating 2a of above-mentioned optical-waveguide sensor chip.

Below, explanation uses above-mentioned optical waveguide sensor to carry out the method for substance-measuring with reference to Fig. 4 (A)～(C).

At first, the optical-waveguide sensor chip shown in the set-up dirgram 4 (A).This sensor chip comprises the substrate 1 with grating 2a, 2b.Planar optical waveguide 3 forms on substrate 1 principal plane that comprises grating 2a, 2b.Low refractive index resin 4 covers on the planar optical waveguide 3, and according to the mode opening that a part that makes the planar optical waveguide 3 between grating 2a, 2b is exposed, for example forms rectangular reacting hole 5.Be immobilized in from planar optical waveguide 3 surfaces that reacting hole 5 exposes with the 1st material (for example the 1st antibody) 11 of determination object material (for example antigen) specific reaction of determined sample.Back up pad 7 forms according to the mode that covers reacting hole 5 on low refractive index resin film 4.Immobilization has a plurality of particulates 13 with the 2nd material (for example the 2nd antibody) 12 of the determination object material specific reaction of determined sample to be dispersed on back up pad 7 lower surfaces that reacting hole 5 exposes.

Then, the dropping via determined sample solution drips determined sample solution with hole (not shown) in reacting hole 5.At this moment, if do not exist in the determined sample solution that drips with the 1st antibody 11 on planar optical waveguide 3 surfaces with the 2nd antibody 12 of particulate 13 antigen of specific reactions take place, then shown in Fig. 4 (B), the 2nd antibody 12 of particulate 13 does not combine with the 1st antibody 11 on planar optical waveguide 3 surfaces, but is dispersed in the determined sample solution 14.Under this state, make red laser incide planar optical waveguide 3 from light incident side grating 2a from red laser diode 21, near surface (exposing surfaces at reacting hole 5 places), send the light that dies in wink even in this planar optical waveguide 3, conduct, because particulate 13 is dispersed in the determined sample solution 14, so particulate 13 exists in the light zone that dies in wink hardly.That is to say that particulate 13 is irrelevant substantially with the absorption or the scattering of the light that dies in wink, the decay of the light intensity that therefore takes place hardly to die in wink.Consequently, the red laser that penetrates from emitting side grating 2b receives the light time at photodiode 22, and its laser light intensity does not almost change.

On the other hand, if have antigen in the determined sample solution 14 that drips, then shown in Fig. 4 (C), antigen-antibody reactions take place with the 1st antibody 11 on planar optical waveguide 3 surfaces and combine in antigen 15, and then the 2nd antibody 12 of particulate 13 is with antigen 15 generation antigen-antibody reactions and combine.That is, between the 2nd antibody 12 of the 1st antibody 11 on planar optical waveguide 3 surfaces and particulate 13, be situated between and antigen-antibody reactions take place, so particulate 13 is immobilized on planar optical waveguide 3 surfaces by antigen 15.

After above-mentioned determined sample solution just drips, make red laser incide planar optical waveguide 3 from light incident side grating 2a from red laser diode 21, near surface (exposing surfaces at reacting hole 5 places), send the light that dies in wink if in this planar optical waveguide 3, conduct, then be immobilized on planar optical waveguide 3 surfaces, so particulate 13 exists at the light region memory that dies in wink owing to particulate 13.That is to say that particulate 13 is relevant with the absorption or the scattering of the light that dies in wink, the decay of the light intensity that therefore takes place to die in wink.Consequently, the red laser that penetrates from emitting side grating 2b receives the light time at photodiode 22, and its laser light intensity is because the influence of immobilized particulate 13 and along with the time reduces.

The reduced rate of the laser light intensity that receives at photodiode 22 places and the amount of the particulate 13 that is immobilized onto planar optical waveguide 3 surfaces, promptly the antigen concentration in the relevant determined sample solution 14 is proportional with antigen-antibody reaction.Therefore, be produced in the known determined sample solution of antigen concentration the reduction curve of laser light intensity as time passes, obtain the reduced rate of the laser light intensity that this curve locates at the appointed time, make the typical curve of the relation between the reduced rate of expression antigen concentration and laser light intensity in advance.From obtaining the reduced rate of laser light intensity at the appointed time with the time of said method mensuration and the reduction curve of laser light intensity, with the reduced rate of this laser light intensity and the contrast of above-mentioned typical curve, thereby can measure antigen concentration in the determined sample solution.

In addition, in above-mentioned concentration determination, be dispersed on the planar optical waveguide jointly by making particulate and water-soluble substances, thus the dispersiveness of raising particulate.In addition, when planar optical waveguide dripped determined sample solution, the water-soluble substances dissolving with particulate coexistence made particulate to move, thereby the reaction of the 2nd material of determination object material in the determined sample solution and particulate is carried out smoothly.

More than, according to the 2nd embodiment, can provide minimum essential determined sample size few (for example 10 μ L are following), and can carry out quantitative optical-waveguide sensor chip, its manufacture method and optical-waveguide sensor to the concentration of the determination object material of determined sample by determined sample drop being added to the single job of measuring the district.

In addition, according to the 2nd embodiment, can provide minimum essential determined sample size few (for example 10 μ L are following), and can carry out the assay method of quantitative material to the concentration of the determination object material of determined sample by determined sample drop being added to the single job of measuring the district.

(the 3rd embodiment)

The assay method of the material of the 3rd embodiment below is described.

At first, prepare optical-waveguide sensor chip, it has immobilization from the teeth outwards optical waveguide with the 1st material of determination object material specific reaction is arranged.Then, drip determined sample solution, make between the 1st material of light guide surface and the determination object material in the determined sample solution specific reaction takes place to light guide surface.Then, washing light guide surface.Then, dripping immobilization to light guide surface has dispersion liquid with the particulate of the 2nd material of said determination object material specific reaction, makes between the 2nd material of the determination object material of determined sample solution and particulate specific reaction takes place; By detect be situated between by 1st material and determination object material be immobilized onto the optical change that particulate produced of light guide surface, thereby measure the concentration of determination object material in determined sample solution thereafter.

Optical waveguide for example can be used planar optical waveguide.What illustrate in this planar optical waveguide and the 1st embodiment is same, can be formed by heat-curing resin or alkali-free glass.Will with the 1st material of the determination object material specific reaction of determined sample with the same method immobilization that illustrates in the 1st embodiment on planar optical waveguide.The 1st material is under the situation of antigen at the determination object material of determined sample for example, can use antibody.

Washing for example can use the cleansing solution that has the solution of damping fluid and surfactant etc., the phosphate buffer normal saline (PBS) that contains surfactant, Tris-hydrochloride buffer physiological saline, Good buffer buffer saline, phosphate buffer etc. to form by combination to carry out.

What illustrate in particulate and the 1st embodiment is same, for example can use the inorganic oxide particle of the metallic colloid of the resin beads of LATEX BEAD etc. or collaurum etc. or Titanium particles etc. etc.Particulate also can use the nonmetal particle of polysaccharide, silicon dioxide granule, carbon particle of protein, agarose of albumin etc. etc. etc.Preferred especially LATEX BEAD, metallic colloid.Particulate preferably has the diameter of 50nm～10 μ m.

With the 2nd material use with the 1st embodiment in the same method immobilization that illustrates to particulate.The 2nd material is under the situation of antigen at the determination object material of determined sample for example, can use antibody.

The dispersion liquid of particulate comprises: in damping fluid that contains for example phosphoric acid, trishydroxymethylaminomethane, boric acid, acetate, citric acid, carbonic acid etc. or Good buffer, add the solution that non-ionic surfactants such as stabilizing agent, Tween, Triton-X such as bovine serum albumin(BSA) (BSA), casein, polyglycol form, or phosphate buffer normal saline (PBS) etc.

The assay method that the optical-waveguide sensor chip that reference is shown in Figure 5 and Fig. 6 (A)～(C) specify the material of the 3rd embodiment.

At first, the optical-waveguide sensor chip shown in the set-up dirgram 5 and 6 (A).The dispersion layer of the particulate of Fig. 1 that this sensor chip illustrates, has same structure in not having the 1st embodiment.That is to say, comprise substrate 1 with grating 2a, 2b.Planar optical waveguide 3 forms on substrate 1 principal plane that comprises grating 2a, 2b.Low refractive index resin 4 covers on the planar optical waveguide 3 and according to the mode opening that a part that makes the planar optical waveguide 3 between grating 2a, 2b is exposed, for example forms rectangular reacting hole 5.Be immobilized in from planar optical waveguide 3 surfaces that reacting hole 5 exposes with the 1st material (for example the 1st antibody) 11 of determination object material (for example antigen) specific reaction of determined sample.In Fig. 5, variation for the light that dies in wink of measuring planar optical waveguide 3 places that are positioned at reacting hole 5, setting makes light incide the laser oscillator (for example red laser diode) 21 of light incident side grating 2a, and the photo-electric conversion element (photodiode) 22 of reception from the light of emitting side grating 2b ejaculation is set.

Then, in reacting hole 5, drip determined sample solution.At this moment, shown in Fig. 6 (B), antigen-antibody reaction takes place with the 1st antibody 11 on planar optical waveguide 3 surfaces and combines in the antigen 15 in the determined sample solution of dropping.

Then, carry out carrying out washing treatment to rinse out not the antigen 15 with the 1st antibody 11 reactions on planar optical waveguide 3 surfaces.Then, dripping in reacting hole 5 that immobilization has with the determination object material is the dispersion liquid of particulate of the 2nd material (for example the 2nd antibody) of antigentic specificity reaction.At this moment, shown in Fig. 6 (C), in dispersion liquid 16, be antigen 15 with the 2nd antibody 12 generation antigen-antibody reactions of particulate 13 with antigen-antibody reactions take place the 1st antibody 11 on planar optical waveguide 3 surfaces determination object material and combined.That is to say, be situated between between the 1st antibody 11 on planar optical waveguide 3 surfaces and the 2nd antibody 12 of particulate 13 and antigen-antibody reaction take place, thereby particulate 13 is immobilized planar optical waveguide 3 surfaces by antigen 15.

After the dispersion liquid of particulate has just dripped, make red laser incide planar optical waveguide 3 from light incident side grating 2a from red laser diode 21, if in 3 conduction of this planar optical waveguide and near surface (exposing surfaces at reacting hole 5 places), send the light that dies in wink, then be immobilized on planar optical waveguide 3 surfaces, so particulate 13 exists at the light region memory that dies in wink owing to particulate 13.That is to say that particulate 13 is relevant with the absorption or the scattering of the light that dies in wink, the decay of the light intensity that therefore takes place to die in wink.Consequently, the red laser that penetrates from emitting side grating 2b receives the light time at photodiode 22, and its laser light intensity is because the influence of immobilized particulate 13 and along with the time reduces.

The reduced rate of the laser light intensity that receives at photodiode 22 places and the amount of the particulate 13 that is immobilized onto planar optical waveguide 3 surfaces, promptly the antigen concentration in the relevant determined sample solution 14 is proportional with antigen-antibody reaction.Therefore, be produced in the known determined sample solution of antigen concentration the reduction curve of laser light intensity as time passes, obtain the reduced rate of the laser light intensity that this curve locates at the appointed time, make the typical curve of the relation between the reduced rate of expression antigen concentration and laser light intensity in advance.Obtain the reduced rate of laser light intensity at the appointed time from the reduction curve of the time measured with said method and laser light intensity, with the reduced rate of this laser light intensity and the contrast of above-mentioned typical curve, thereby can measure antigen concentration in the determined sample solution.

More than, according to the 3rd embodiment, can provide minimum essential determined sample size few (for example 10 μ L are following), and be added drop-wise to by determined sample solution and to measure district, washing and particle dispersion liquid and be added drop-wise to the assay method that 3 operations measuring the district can be carried out the determination object thing of quantitative determined sample to the concentration of the determination object material of determined sample.

Particularly, in the 3rd embodiment, washing after measuring the district because determined sample solution is added drop-wise to, under the high situation of the concentration of the determination object material (for example antigen) of determined sample solution, is effective therefore.

(the 4th embodiment)

The assay method of the material of the 4th embodiment below is described.

At first, prepare optical-waveguide sensor chip, it has immobilization from the teeth outwards optical waveguide with the 1st material of determination object material specific reaction is arranged.At first, determined sample solution and immobilization had with the particulate of the 2nd material of the determination object material specific reaction of determined sample in the container of for example microtubule, mix, make between the determination object material of the 2nd material of particulate and determined sample solution specific reaction takes place.Then, above-mentioned mixed liquor is added drop-wise to the light guide surface of the sensor chip, make the 1st material of light guide surface with the determination object material generation specific reaction of the determined sample solution of the 2nd substance reaction of particulate.Then, be immobilized onto the particulate of light guide surface by the 1st material and determination object material, promptly be immobilized onto the optical change that particulate produced of light guide surface, thereby measure the concentration of the determination object material in the determined sample solution by detecting to be situated between.

Optical waveguide for example can be used planar optical waveguide.What illustrate in this planar optical waveguide and the 1st embodiment is same, can be formed by heat-curing resin or alkali-free glass.Will with the 1st material of the determination object material specific reaction of determined sample with the same method immobilization that illustrates in the 1st embodiment on planar optical waveguide.The 1st material is under the situation of antigen at the determination object material of determined sample for example, can use antibody.

What illustrate in particulate and the 1st embodiment is same, for example can use the inorganic oxide particle of the metallic colloid of the resin beads of LATEX BEAD etc. or collaurum etc. or Titanium particles etc. etc., preferred especially LATEX BEAD, metallic colloid.

Particulate preferably has the diameter of 50nm～10 μ m.

With the 2nd material use with the 1st embodiment in the same method immobilization that illustrates to particulate.The 2nd material is under the situation of antigen at the determination object material of determined sample for example, can use antibody.

Microparticulate is added the solution that non-ionic surfactants such as stabilizing agent, Tween, Triton-X such as bovine serum albumin(BSA) (BSA), casein, polyglycol form in damping fluid that is containing for example phosphoric acid, trishydroxymethylaminomethane, boric acid, acetate, citric acid, carbonic acid etc. or Good buffer, perhaps in phosphate buffer normal saline (PBS) etc., can prepare the dispersion liquid of particulate.

Determined sample solution and immobilization have with the mixing of the particulate of the 2nd material of the determination object material specific reaction of determined sample in, particulate can be the state of dispersion liquid, also can be the state of solid (for example dry substance, scars or powder).Particularly, prepare the dispersion liquid of particulate in advance, this dispersion liquid mixes in the container of for example microtubule with determined sample solution then.In addition, when having the particulate of the 2nd material in the container of for example microtubule, to mix determined sample solution and immobilization, dispersion liquid and dry, for example freeze drying that also can in container, add the particulate that contains water-soluble substances earlier, particulate is distributed to inner surface of container by water-soluble substances, in container, adds determined sample solution and mixing thereafter.Water-soluble substances for example can use polyvinyl alcohol (PVA), bovine serum albumin(BSA) (BSA), polyglycol, phospholipid polyalcohol, gelatin, carbohydrate (for example sucrose, trehalose).

The assay method that specifies the 4th embodiment with reference to above-mentioned optical-waveguide sensor chip shown in Figure 5 and Fig. 7 (A), (B).

At first, shown in Fig. 5 and 7 (A), preparing on planar optical waveguide 3 surfaces of exposing from reacting hole 5 immobilization has optical-waveguide sensor with the 1st material (for example the 1st antibody) 11 of determination object material (for example antigen) specific reaction of determined sample.

At first, having determined sample solution and immobilization with the determination object material is that the dispersion liquid of particulate of the 2nd material (for example the 2nd antibody) of antigentic specificity reaction is for example mixing in the microtubule, makes the 2nd antibody generation antigen-antibody reaction of the antigen and the particulate of determined sample solution.

Then, in reacting hole 5, drip above-mentioned mixed liquor.At this moment, shown in Fig. 7 (B), in the mixed liquor with the determined sample solution of the 2nd antibody generation antigen-antibody reaction in antigen 15 with the 1st antibody 11 on planar optical waveguide surface antigen-antibody reactions take place and combine.That is to say that the antigen 15 that has combined with the 2nd antibody 12 of particulate 13 with the 1st antibody 11 on planar optical waveguide 3 surfaces antigen-antibody reaction takes place and combines in advance, consequently particulate 13 is immobilized planar optical waveguide 3 surfaces.

After the dispersion liquid of above-mentioned determined sample solution and particulate has just dripped, make red laser incide planar optical waveguide 3 from light incident side grating 2a from red laser diode 21, near surface (exposing surfaces at reacting hole 5 places), send the light that dies in wink if in this planar optical waveguide 3, conduct, then be immobilized on planar optical waveguide 3 surfaces, so particulate 13 exists at the light region memory that dies in wink owing to particulate 13.That is to say that particulate 13 is relevant with the absorption or the scattering of the light that dies in wink, therefore cause the decay of the light intensity that dies in wink.Consequently, the red laser that penetrates from emitting side grating 2b receives the light time at photodiode 22, and its laser light intensity is because the influence of immobilized particulate 13 and along with the time reduces.

The reduced rate of the laser light intensity that receives at photodiode 22 places and the amount of the particulate 13 that is immobilized onto planar optical waveguide 3 surfaces, promptly the antigen concentration in the relevant determined sample solution 14 is proportional with antigen-antibody reaction.Therefore, be produced in the known determined sample solution of antigen concentration the reduction curve of laser light intensity as time passes, obtain the reduced rate of the laser light intensity that this curve locates at the appointed time, make the typical curve of the relation between the reduced rate of expression antigen concentration and laser light intensity in advance.Obtain the reduced rate of laser light intensity at the appointed time from the reduction curve of the time measured with said method and laser light intensity, with the reduced rate of this laser light intensity and the contrast of above-mentioned typical curve, thereby can measure antigen concentration in the determined sample solution.

More than, according to the 4th embodiment, can provide minimum essential determined sample size few (for example 10 μ L are following), and can carry out the assay method of the determination object thing of quantitative determined sample to the concentration of the determination object material of determined sample by determined sample solution and particle dispersion liquid being added drop-wise to 1 operation measuring the district.

In addition, in above-mentioned the 4th embodiment, be added dropwise to the determined sample solution of light guide surface and the dispersion liquid of particulate and be pre-mixed, as an alternative, also the dispersion liquid of determined sample solution and particulate can be added drop-wise to light guide surface simultaneously.

In above-mentioned the 4th embodiment, be added drop-wise to light guide surface after the dispersion liquid mixing with determined sample solution and particulate, but also can be to drip the order that drips the dispersion liquid of particulate behind the determined sample solution, also can be to drip the order that drips determined sample solution behind the dispersion liquid of particulate.No matter be assay method with the material of which kind of order, all same with the 4th embodiment, can few with the essential determined sample size of minimum (for example 10 μ L are following) concentration of the determination object material of determined sample be carried out quantitatively.

(the 5th embodiment)

The following describes the substance-measuring kit of the 5th embodiment.

Substance-measuring kit is made of combination optical-waveguide sensor chip and package body: above-mentioned package body is taken in immobilization dispersion liquid with the particulate of the 2nd material of the determination object material specific reaction of determined sample.Optical-waveguide sensor chip has optical waveguide and lid, described optical waveguide immobilization from the teeth outwards has the 1st material with the determination object material specific reaction of determined sample, described lid is disposed on this optical waveguide, have the recess that is used for forming opposite to each other the mensuration district, and opening there be entrance hole and the exhaust opening that is communicated with this mensuration district with optical waveguide.

Will with the 1st material of the determination object material specific reaction of determined sample use with the 1st embodiment in the same method immobilization that illustrates to planar optical waveguide.The 1st material is under the situation of antigen at the determination object material of determined sample for example, can use antibody.

Be accommodated in illustrate in the particulate of package body and the 1st embodiment same, for example can use the inorganic oxide particle of the metallic colloid of the resin beads of LATEX BEAD etc. of POLYSTYRENE system or collaurum etc. or Titanium particles etc. etc.Particulate also can use the nonmetal particle of polysaccharide, silicon dioxide granule, carbon particle of protein, agarose of albumin etc. etc. etc.Preferred especially LATEX BEAD, metallic colloid.Particulate preferably has the diameter of 50nm～10 μ m.

With the 2nd material use with the 1st embodiment in the same method immobilization that illustrates to particulate.The 2nd material is under the situation of antigen at the determination object material of determined sample for example, can use antibody.

The dispersion liquid of particulate comprises: add the solution that non-ionic surfactants such as stabilizing agent, Tween, Triton-X such as bovine serum albumin(BSA) (BSA), casein, polyglycol form in damping fluid that contains for example phosphoric acid, trishydroxymethylaminomethane, boric acid, acetate, citric acid, carbonic acid etc. or Good buffer, perhaps phosphate buffer normal saline (PBS) etc.

Package body can be made by the stacked film of for example polyethylene film or tygon and polyethylene terephthalate.In addition, package body can use microtubule, plastic bottle, vial.

Specify the substance-measuring kit of the 5th embodiment with reference to the (a) and (b) of above-mentioned Fig. 8.Fig. 8 (a) is the vertical view of expression optical-waveguide sensor chip, and figure (b) is the sectional view of above-mentioned figure (a).

Both ends at the principal plane of glass substrate 31 are provided with light incident side grating 32a and the emitting side grating 32b that is for example formed by titanium dioxide.For example the planar optical waveguide 33 that is made of heat-curing resin forms on substrate 31 principal planes that comprise grating 32a, 32b.The lid of being made by for example acryl resin 34 disposes according to the principal plane of overlay planes optical waveguide 33 and the mode of side.In addition, here, the also available replacements such as other resin of material of lid 34 with low-refraction of regulation.The lid 34 and planar optical waveguide 33 surfaces between have the rectangle recess 36 that is used to form for example rectangular mensuration district 35.In addition, covering 34 upper sheds has from its surface to entrance hole 37 and the exhaust opening 38 of measuring district 35.Handle by the hydrophobization that adopts silane coupling agent with the 1st material 11 of the determination object material specific reaction of determined sample and immobilization to from measuring on 35 planar optical waveguides of exposing, 33 surfaces, district.Constitute optical-waveguide sensor chip by such planar optical waveguide 33, lid 34 etc.

Immobilization has the dispersion liquid with the particulate of the 2nd material of the determination object material specific reaction of determined sample for example to be incorporated in the package body (not shown) of tygon system, constitutes to measure with above-mentioned optical-waveguide sensor chip combination and uses kit.

Below, explanation uses the mentioned reagent box to carry out the method for substance-measuring with reference to Fig. 9 (A)～(C).In addition, in order to measure from the variation of the light that dies in wink of measuring the planar optical waveguide of exposing in the district, setting makes light incide the laser oscillator (for example red laser diode) 21 of light incident side grating 32a, and the photoelectric conversion element (photodiode) 22 of reception from the light of emitting side grating 32b ejaculation is set.

At first, shown in Fig. 8 and Fig. 9 (A), preparing is having optical-waveguide sensor chip with the 1st material (for example the 1st antibody) 11 of determination object material (for example antigen) specific reaction of determined sample from measuring immobilization on 35 planar optical waveguides of exposing, 33 surfaces, district.

Then, in measuring district 35, drip determined sample solution via entrance hole 37.At this moment, shown in Fig. 9 (B), antigen-antibody reaction takes place with the 1st antibody 11 on planar optical waveguide 33 surfaces and combines in the antigen 15 in the determined sample solution 14 of dropping.

Then, the dispersion liquid of the particulate in the package body is imported to planar optical waveguide 33 surfaces of measuring in the district 35 via the entrance hole 37 of lid 34, simultaneously determined sample solution is discharged to the outside via exhaust opening 38.In the meantime, unreacted antigen and the dispersion liquid in the determined sample liquid together is rinsed.Simultaneously, shown in Fig. 9 (C), the 2nd antibody 12 of the particulate 13 in the dispersion liquid 16 be that antigen 15 antigen-antibody reactions take place and combines with antigen-antibody reactions take place the 1st antibody 11 on planar optical waveguide 33 surfaces determination object material.That is to say, be situated between between the 1st antibody 11 on planar optical waveguide 33 surfaces and the 2nd antibody of particulate 13 and antigen-antibody reaction take place, thereby make particulate 13 immobilizations to planar optical waveguide 33 surfaces by antigen 15.

After the dispersion liquid of above-mentioned particulate has just imported, make red laser incide planar optical waveguide 33 from light incident side grating 32a from red laser diode 21, near conduction and the surface exposing surfaces of 35 places (measure district), sending under the situation of the light that dies in wink in this planar optical waveguide 33, because the particulate 13 in the dispersion liquid 16 is immobilized on planar optical waveguide 33 surfaces, so particulate 13 exists at the light region memory that dies in wink.That is to say that particulate 13 is relevant with the absorption or the scattering of the light that dies in wink, therefore cause the decay of the light intensity that dies in wink.Consequently, the red laser that penetrates from emitting side grating 32b receives the light time at photodiode 22, and its laser light intensity is because the influence of immobilized particulate 13 and along with the time reduces.

The amount of the particulate 13 that the reduced rate of the laser light intensity that receives at photodiode 22 places and planar optical waveguide 33 are surface immobilized, promptly the antigen concentration in the relevant determined sample solution 14 is proportional with antigen-antibody reaction.Therefore, be produced in the known determined sample solution of antigen concentration the reduction curve of laser light intensity as time passes, obtain the reduced rate of the laser light intensity that this curve locates at the appointed time, make the typical curve of the relation between the reduced rate of expression antigen concentration and laser light intensity in advance.Obtain the reduced rate of laser light intensity at the appointed time from the reduction curve of the time measured with said method and laser light intensity, with the reduced rate of this laser light intensity and the contrast of above-mentioned typical curve, thereby can measure antigen concentration in the determined sample solution.

More than, substance-measuring kit according to the 5th embodiment, owing to the following parts of combination constitute: optical waveguide and drip determined sample solution and importing to measuring the district, the optical-waveguide sensor chip of the structure that the dispersion liquid of discharge particulate can carry out, and the package body of taking in particle dispersion liquid, therefore can few with the essential determined sample size of minimum (for example 10 μ L be following), and by determined sample solution is added drop-wise to measure the district and with the dispersion liquid of particulate to measuring that the district imports and from measure 2 operations of discharging in the district, can carry out quantitatively the concentration of the determination object material of determined sample.

Particularly, in the assay method of the determination object thing of the determined sample that uses substance-measuring kit to carry out, in that being added drop-wise to, determined sample solution measures the laggard dispersion liquid of being about to particulate in district to measuring that the district imports and discharging from measuring the district, therefore under the high situation of the concentration of the determination object material (for example antigen) of determined sample solution, also be effective.

Below, describe embodiments of the invention in detail with reference to above-mentioned accompanying drawing.

(embodiment 1)

With refractive index be 2.2～2.4 titanium dioxide to be splashed to refractive index be that forming thickness is the oxidation titanium film of 50nm on 1.52 the alkali-free glass substrate, utilize then photoetching process and dried quarter method on glass substrate 1, form grating 2a, 2b.Then, epoxy resin solution is spun on the glass substrate 1 that comprises grating 2a, 2b, forms the planar optical waveguide 3 that thickness is about 30 μ m by calcining then.The refractive index of the planar optical waveguide 3 after the calcining is 1.56.Then, by with poly-(perfluorobuttene base vinyl ether) screen printing of the CYTOP (registered trademark) of low refractive index resin, commercially available Asahi Glass Co., Ltd's system on planar optical waveguide 3, form the low refractive index resin 4 that opening has rectangular reacting hole (measuring the district) 5.

Then, utilize the silane coupling agent will be surface-hydrophobicized from the planar optical waveguide 3 that reacting hole 5 exposes, by hydrophobic interaction with anti-insulin antibody 11 immobilizations on it.On low refractive index resin 4 according to the mode of surrounding reacting hole 5 form the cell wall 6 of frame shape thereafter.

In addition, Blocking One (Nacalai Tesque Co., Ltd. system) is joined in the phosphate buffer normal saline (PBS) according to the mode that is diluted to 2.5 times, with preparation solution.Then, it is that the blue LATEX BEAD of 760nm is distributed in this solution that immobilization is had the mean grain size of anti-insulin antibody, is the bead dispersion liquid of 4 weight % thereby the preparation bead disperses concentration.Wherein, BlockingOne is the sealer that is used to suppress non-specific adsorption, and it is the aqueous solution of the 4M-sodium hydroxide solution of the following antiseptic of albumin, the casein of 2～6 weight %, the 10 weight % of three (methylol) aminomethane of containing 4～8 weight %, 1～2 weight % following macromolecular compound, 1 weight % and about 3 weight %.

There is the bead dispersant liquid drop of anti-insulin antibody to be added in the above-mentioned reacting hole 5 10 these immobilizations of μ L,, about 1 day then, makes the optical-waveguide sensor chip as shown in Figure 1 that is provided with bead in advance by freeze drying at-80 ℃ of following precoolings.When this freeze drying, in the bead dispersion liquid of above-mentioned composition, add disaccharides trehalose 3 weight %, surfactant Tween 0.1 weight %.The purpose of adding these compositions is to improve the redispersibility of bead dispersion liquid.

Dripping determined sample solution in the reacting hole of the sensor chip that obtains respectively is that concentration is the insulin solutions of 1.6ng/mL and 6.4ng/mL, makes it to take place antigen-antibody reaction.After determined sample solution just drips, the red light that will be 655nm from the wavelength of red LED 21 incides planar optical waveguide 3 via light incident side grating 2a, in this planar optical waveguide 3 conduction takes place and light takes place to die in wink near surface (exposing surfaces at reacting hole 5 places), receive the red light that penetrates from emitting side grating 2b at photodiode 22 places, measure its light intensity.That is to say, measure the variation of light intensity as time passes.

For the insulin solutions of 6.4ng/mL, carry out 3 (amounting to 4 times) same operations, measure the variation of light intensity as time passes.These the results are shown in Figure 10.

In Figure 10, be 100% with the light intensity after the dropping just of determined sample solution, represent the variation of its light intensity as time passes.Among Figure 10, concentration is that the result of the insulin solutions of 1.6ng/mL is expressed as S1, and concentration is that the result of 4 insulin solutions of 6.4ng/mL is expressed as S2-1, S2-2, S2-3, S2-4.In addition, use only for the diluting solvent of insulin carries out same operation as determined sample solution (blank), the result of the variation of the laser light intensity as time passes of mensuration is expressed as B in Figure 10.

As can be seen from Figure 10, the reduced rate of the laser light intensity at stipulated time place is relevant with insulin concentration in the determined sample solution.In addition as can be known, in 4 determined sample solutions of insulin concentration identical (6.4ng/mL), the reduced rate of the laser light intensity that the stipulated time is located is approximate, therefore can carry out the concentration determination of favorable reproducibility.

(embodiment 2)

With refractive index be 2.2～2.4 titanium dioxide to be splashed to refractive index be that forming thickness is the oxidation titanium film of 50nm on 1.52 the alkali-free glass substrate 1, utilize then photoetching process and dried quarter method on glass substrate 1, form grating 2a, 2b.Then, epoxy resin solution is spun on the glass substrate 1 that comprises grating 2a, 2b, forms the planar optical waveguide 3 that thickness is about 30 μ m by calcining then.The refractive index of the planar optical waveguide 3 after the calcining is 1.56.Then, by with poly-(perfluorobuttene base vinyl ether) screen printing of the CYTOP (registered trademark) of low refractive index resin, commercially available Asahi Glass Co., Ltd's system on planar optical waveguide 3, forming opening has the low refractive index resin 4 of rectangular reacting hole (measuring the district) 5, thereby makes optical-waveguide sensor chip as shown in Figure 5.

Dripping determined sample solution in the reacting hole of the sensor chip that obtains respectively is that concentration is the insulin solutions 10 μ L of 1.6ng/mL and 6.4ng/mL, carries out antigen-antibody reaction in 10 minutes under 37 ℃.Then, use remaining insulin in the lavation buffer solution washing reaction hole of forming by Tris buffer saline (TBS).Drip bead dispersion liquid 20 μ L similarly to Example 1 in the reacting hole after washing.When the bead dispersion liquid has just dripped, measure the variation of light intensities similarly to Example 1 with red LED 21 and photodiode 22.

For the insulin solutions of 6.4ng/mL, carry out 3 (amounting to 4 times) same operations, measure the variation of light intensity as time passes.With the light intensity after the dropping just of determined sample solution is 100%, and the variation of its light intensity as time passes as shown in figure 11.Among Figure 11, concentration is that the result of the insulin solutions of 1.6ng/mL is expressed as S1, and concentration is that the result of 4 insulin solutions of 6.4ng/mL is expressed as S2-1, S2-2, S2-3, S2-4.In addition, use only for the diluting solvent of insulin carries out same operation as determined sample solution (blank), the result of the variation of the laser light intensity as time passes of mensuration is expressed as B in Figure 11.

As can be seen from Figure 11, the reduced rate of the laser light intensity at stipulated time place is relevant with insulin concentration in the determined sample solution.In addition as can be known, in 4 determined sample solutions of insulin concentration identical (6.4ng/mL), the reduced rate of the laser light intensity that the stipulated time is located is approximate, therefore can carry out the concentration determination of favorable reproducibility.

(embodiment 3)

In the reacting hole of similarly to Example 2 optical-waveguide sensor chip, drip bead dispersion liquid 10 μ L similarly to Example 1, thereafter dripping determined sample solution immediately respectively is that concentration is the insulin solutions 10 μ L of 1.6ng/mL and 6.4ng/mL, stirs by imbibition.Immediately, measure the variation of light intensities similarly to Example 1 with red LED 21 and photodiode 22 after stir.

For the insulin solutions of 6.4ng/mL, carry out 3 (amounting to 4 times) same operations, measure the variation of light intensity as time passes.With the light intensity after the dropping just of determined sample solution is 100%, and the variation of its light intensity as time passes as shown in figure 12.Among Figure 12, concentration is that the result of the insulin solutions of 1.6ng/mL is expressed as S1, and concentration is that the result of 4 insulin solutions of 6.4ng/mL is expressed as S2-1, S2-2, S2-3, S2-4.In addition, use only for the diluting solvent of insulin carries out same operation as determined sample solution (blank), the result of the variation of the laser light intensity as time passes of mensuration is expressed as B in Figure 12.

As can be seen from Figure 12, the reduced rate of the laser light intensity at stipulated time place is relevant with insulin concentration in the determined sample solution.In addition as can be known, in 4 determined sample solutions of insulin concentration identical (6.4ng/mL), the reduced rate of the laser light intensity that the stipulated time is located is approximate, therefore can carry out the concentration determination of favorable reproducibility.

In addition, in embodiment 3,, can confirm similarly to Example 3 that all the reduced rate of laser light intensity is relevant with insulin concentration in the determined sample solution even the intermodulation of the dropping of bead dispersion liquid and insulin solutions order or drips simultaneously.

(embodiment 4)

At first, with also freeze drying in the bead dispersion liquid 50 μ L adding microtubule similarly to Example 1.When freeze drying, in above-mentioned bead dispersion liquid, add disaccharides trehalose 3 weight %, surfactant Tween 0.1 weight % similarly to Example 1.Then, dripping determined sample solution in microtubule respectively is that concentration is the insulin solutions 50 μ L of 1.6ng/mL and 6.4ng/mL, mixes and carries out antigen-antibody reaction.Then, mixed liquor in the microtubule 20 μ L are added drop-wise in the reacting hole of optical-waveguide sensor chip as shown in Figure 5 similarly to Example 2, after dropping, begin to measure the variation of light intensity immediately similarly to Example 1 with red LED 21 and photodiode 22.

For the insulin solutions of 6.4ng/mL, carry out 3 (amounting to 4 times) same operations, measure the variation of light intensity as time passes.With the light intensity after the dropping just of determined sample solution is 100%, and the variation of its light intensity as time passes as shown in figure 13.Among Figure 13, concentration is that the result of the insulin solutions of 1.6ng/mL is expressed as S1, and concentration is that the result of 4 insulin solutions of 6.4ng/mL is expressed as S2-1, S2-2, S2-3, S2-4.In addition, use only for the diluting solvent of insulin carries out same operation as determined sample solution (blank), the result of the variation of the laser light intensity as time passes of mensuration is expressed as B in Figure 13.

As can be seen from Figure 13, the reduced rate of the laser light intensity at stipulated time place is relevant with insulin concentration in the determined sample solution.In addition as can be known, in 4 determined sample solutions of insulin concentration identical (6.4ng/mL), the reduced rate of the laser light intensity that the stipulated time is located is approximate, therefore can carry out the concentration determination of favorable reproducibility.

Claims (23)

1. optical-waveguide sensor chip is characterized in that it has:

Immobilization has the optical waveguide with the 1st material of determination object material specific reaction from the teeth outwards; With,

Be dispersed on the described optical waveguide and immobilization has particulate with the 2nd material of described determination object material specific reaction.

2. optical-waveguide sensor chip is characterized in that it has:

Immobilization has the optical waveguide with the 1st material of determination object material specific reaction from the teeth outwards;

The back up pad that disposes in opposite directions with described optical waveguide; With,

Be dispersed in described back up pad with described optical waveguide surface in opposite directions on and immobilization particulate with the 2nd material of described determination object material specific reaction is arranged.

3. optical-waveguide sensor chip as claimed in claim 1 or 2 is characterized in that, described optical waveguide is the glass of sheet.

4. optical-waveguide sensor chip as claimed in claim 1 or 2 is characterized in that, described optical waveguide is that thickness is organic resin film of 3～300 μ m.

5. optical-waveguide sensor chip as claimed in claim 1 or 2 is characterized in that, described particulate is a resin beads.

6. optical-waveguide sensor chip as claimed in claim 1 or 2 is characterized in that, described particulate is a metallic colloid.

7. optical-waveguide sensor chip as claimed in claim 1 or 2, it is characterized in that, described determination object material is an antigen, is respectively antibody with the 1st, the 2nd material that is immobilized in the described determination object material generation specific reaction on described light guide surface and the described particulate.

8. optical-waveguide sensor chip as claimed in claim 1 is characterized in that, described particulate is situated between and is dispersed on the described light guide surface by confining bed.

9. the manufacture method of an optical-waveguide sensor chip is characterized in that, comprises the steps:

Will be with determination object material specific reaction the 1st substance fixed in light guide surface;

To contain immobilization has with the slurry of the particulate of the 2nd material of described determination object material specific reaction and coats on the described optical waveguide; With,

Dry in described coating back, with described microparticulate on described optical waveguide.

10. the manufacture method of an optical-waveguide sensor chip is characterized in that, comprises the steps:

Will be with determination object material specific reaction the 1st substance fixed in light guide surface;

To contain immobilization has with the slurry of the particulate of the 2nd material of described determination object material specific reaction and coats the back up pad surface;

Dry in described coating back, with described microparticulate on described back up pad; With,

With described back up pad according to its microparticulate face and described optical waveguide mode are in opposite directions disposed spaced apartly.

11. the manufacture method as claim 9 or 10 described optical-waveguide-type chips is characterized in that described slurry contains water-soluble substances.

12. the assay method of a material is characterized in that, comprises the steps:

Prepare optical-waveguide sensor chip, this optical-waveguide sensor chip has: immobilization has with the optical waveguide of the 1st material of determination object material specific reaction and is dispersed on the described optical waveguide and immobilization has particulate with the 2nd material of described determination object material specific reaction from the teeth outwards;

Light guide surface to described sensor chip drips determined sample solution, make between the 1st material of light guide surface and the determination object material in the determined sample solution specific reaction takes place, make described determination object material simultaneously and be dispersed between the 2nd material of the particulate on the described optical waveguide specific reaction takes place; With,

Detect to be situated between and to be immobilized onto the optical change that particulate produced of described light guide surface by the 1st material and determination object material.

13. the assay method of a material is characterized in that, comprises the steps:

Prepare optical-waveguide sensor chip, this optical-waveguide sensor chip has immobilization from the teeth outwards optical waveguide with the 1st material of determination object material specific reaction is arranged;

Light guide surface to described sensor chip drips determined sample solution, makes between the 1st material of light guide surface and the determination object material in the determined sample solution specific reaction takes place;

Wash described light guide surface;

Dripping immobilization to described light guide surface has dispersion liquid with the particulate of the 2nd material of the determination object material specific reaction of determined sample, makes between the 2nd material of the determination object material of determined sample solution and particulate specific reaction takes place; With,

Detect to be situated between and to be immobilized onto the optical change that particulate produced of light guide surface by the 1st material and determination object material.

14. the assay method of a material is characterized in that, comprises the steps:

Prepare optical-waveguide sensor chip, this optical-waveguide sensor chip has immobilization from the teeth outwards optical waveguide with the 1st material of determination object material specific reaction is arranged;

Being pre-mixed determined sample solution and immobilization has particulate with the 2nd material of the determination object material specific reaction of determined sample, makes the 2nd material of particulate and the determination object material generation specific reaction of determined sample solution;

Light guide surface to described sensor chip drips described mixed liquor, make the 1st material of described light guide surface with the determination object material generation specific reaction of the determined sample solution of the 2nd substance reaction of particulate; With,

Detect to be situated between and to be immobilized onto the optical change that particulate produced of described light guide surface by the 1st material and determination object material.

15. the assay method of a material is characterized in that, comprises the steps:

Prepare optical-waveguide sensor chip, this optical-waveguide sensor chip has immobilization from the teeth outwards optical waveguide with the 1st material of determination object material specific reaction is arranged;

Light guide surface to described sensor chip drips determined sample solution, makes between the 1st material of light guide surface and the determination object material in the determined sample solution specific reaction takes place;

Dripping immobilization to described light guide surface has dispersion liquid with the particulate of the 2nd material of the determination object material specific reaction of determined sample, makes between the 2nd material of described determination object material and particulate specific reaction takes place; With,

Detect to be situated between and to be immobilized onto the optical change that particulate produced of described light guide surface by the 1st material and determination object material.

16. the assay method of a material is characterized in that, comprises the steps:

Prepare optical-waveguide sensor chip, this optical-waveguide sensor chip has immobilization from the teeth outwards optical waveguide with the 1st material of determination object material specific reaction is arranged;

Drip immobilization to the light guide surface of described sensor chip dispersion liquid with the particulate of the 2nd material of the determination object material specific reaction of determined sample is arranged;

There is the described light guide surface of described dispersion liquid to drip determined sample solution to dropping, make between the 1st material of light guide surface and the determination object material in the determined sample solution specific reaction takes place, make simultaneously between the 2nd material of particulate in described determination object material and the described dispersion liquid specific reaction takes place; With,

Detect to be situated between and to be immobilized onto the optical change that particulate produced of described light guide surface by the 1st material and determination object material.

17. the assay method of a material is characterized in that, comprises the steps:

Prepare optical-waveguide sensor chip, this optical-waveguide sensor chip has: immobilization has the optical waveguide with the 1st material of determination object material specific reaction from the teeth outwards, the back up pad that disposes in opposite directions with described optical waveguide, and be scattered in described back up pad with surface and the immobilization in opposite directions of described optical waveguide particulate with the 2nd material of described determination object material specific reaction is arranged;

Between the optical waveguide of described sensor chip and described back up pad, inject determined sample solution, make between the 1st material of light guide surface and the determination object material in the determined sample solution specific reaction takes place, make described determination object material simultaneously and be scattered between the 2nd material of particulate of described back up pad specific reaction takes place; With,

Detect to be situated between and to be immobilized onto the optical change that particulate produced of described light guide surface by the 1st material and determination object material.

18. a substance-measuring kit is characterized in that its combination has optical-waveguide sensor chip and package body,

Described optical-waveguide sensor chip has optical waveguide and lid, and described optical waveguide immobilization from the teeth outwards has the 1st material with the determination object material specific reaction of determined sample; Described lid is disposed at described light guide surface, and described light guide surface between have and be used to form the recess of measuring the district, and opening has entrance hole and the exhaust opening that is communicated with this mensuration district;

Described package body is taken in immobilization dispersion liquid with the particulate of the 2nd material of the determination object material specific reaction of determined sample.

19. the assay method of a material is characterized in that, wherein uses the described substance-measuring kit of claim 18, and comprises the steps:

Determined sample solution is added drop-wise to the light guide surface of measuring in the district via the entrance hole of optical-waveguide sensor chip lid, makes the 1st material that is immobilized onto light guide surface and the determination object material generation specific reaction of determined sample solution;

The dispersion liquid of the particulate in the package body is imported to the light guide surface of measuring in the district via the entrance hole of described lid, simultaneously, during determined sample solution is discharged via exhaust opening, make between the 2nd material of the determination object material of the determined sample solution that specific reaction takes place and particulate specific reaction takes place; With

Detect to be situated between and to be immobilized onto the optical change that particulate produced of light guide surface by the 1st material and determination object material.

20. the assay method as claim 12～17 or 19 each described materials is characterized in that, the determination object material of described determined sample solution is an antigen, with described the 1st, the 2nd material of this determination object material generation specific reaction be antibody.

21. substance-measuring kit as claimed in claim 18 is characterized in that, the determination object material of described determined sample solution is an antigen, with described the 1st, the 2nd material of this determination object material generation specific reaction be antibody.

22. an optical-waveguide sensor is characterized in that it has:

Optical-waveguide sensor chip, it has: immobilization has with the optical waveguide of the 1st material of determination object material specific reaction and is scattered on the described optical waveguide and immobilization has particulate with the 2nd material of described determination object material specific reaction from the teeth outwards;

Make light incide the light source of described optical waveguide; With,

Reception is from the photo detector of the light of described optical waveguide ejaculation.

23. an optical-waveguide sensor is characterized in that it has:

Optical-waveguide sensor chip, it has: immobilization has the optical waveguide with the 1st material of determination object material specific reaction from the teeth outwards, the back up pad that disposes in opposite directions with described optical waveguide, and be scattered in described back up pad with surface and the immobilization in opposite directions of described optical waveguide particulate with the 2nd material of described determination object material specific reaction is arranged;

Make light incide the light source of described optical waveguide; With

Reception is from the photo detector of the light of described optical waveguide ejaculation.

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