Chemical apparatus and process disinfecting – deodorizing – preser – Analyzer – structured indicator – or manipulative laboratory... – Means for analyzing liquid or solid sample
Reexamination Certificate
1999-07-06
2002-08-13
Warden, Jill (Department: 1743)
Chemical apparatus and process disinfecting, deodorizing, preser
Analyzer, structured indicator, or manipulative laboratory...
Means for analyzing liquid or solid sample
C422S082050, C422S082060, C422S082080, C422S082090, C385S012000
Reexamination Certificate
active
06432364
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates to an immunoassay apparatus and in particular, to an SPR sensor cell utilizing a phenomenon of surface plasmon resonance (hereinafter, referred to as SPR) and an immunoassay apparatus using the SPR sensor cell.
2. Description of the Related Art
Conventionally, the immunoassay is generally used for detecting a very small quantity of protein. This immunoassay utilizes a specific immune reaction between an antigen (protein to be detected) and a corresponding antibody (prepared using the antigen) so as to determine the antigen concentration in a sample. This immunoassay can determine an antigen even in a sample having a plurality of antigens without isolating the target antigen, unlike a chemical or physical quantity determination.
Moreover, various types of immunoassay are available:
1) radio immunoassay (RIA)
2) enzyme immunoassay (EIA)
3) fluoro immunoassay (FIA)
The RIA (radio immunoassay) needs to use an isotope and is not widely used recently. Moreover, the EIA (enzyme immunoassay) is widely used currently because the EIA can easily determine an immune reaction. Furthermore, the FIA (fluoro immunoassay) can determine a quantity with a higher sensitivity and a higher accuracy.
The EIA method which uses a solid phase for determining a quantity of an antibody is called enzyme-linked immunosorbent assay (ELISA). There are two types of ELISA:
a) indirect method using an antigen in a solid phase; and
b) antibody catching method using anti-IgG antibody in a solid phase.
The ELISA method is used for determining a quantity of antibody for a specific causal organism; determining a quantity of antibody for an allergen; and for screening of a monochronal antibody.
For the ELISA, an assay kit includes a micro-plate having 96 wells on which an immune reaction is determined. Accordingly, it is possible to determine quantities of a multiple samples at once. Recently, various types of the automated ELISA are available on market.
As for the ELISA kit, various chemicals are produced by various companies. For example, tPA is an enzyme which indirectly serves to dissolve fibrin in blood associated with a blood clotting and thrombus. Moreover, PAI-1 suppresses the tPA and serves to make a blood clotting and thrombus.
On the other hand, the so-called SPR sensor is known as a sensor used in an immunoassay apparatus. The SPR sensor is sensor utilizing the surface plasmon resonance phenomenon as follows.
That is, a thin metal film of about 50 nm thickness is deposited onto a bottom of a prism having a high refraction factor. Then, a predetermined light is introduced with an angle greater than a critical angle from the prism to the thin metal film. The thin metal film of 50 nm thickness is semi-transparent. The light incident from the prism passes through the thin metal film to reach a surface of the thin metal film at the opposite side of the prism, generating an evanescent field on the surface of the thin metal film.
By adjusting the light incident angle, the number of waves of the evanescent field is matched with the number of waves of surface plasmon resonance so as to excite a surface plasmon resonance on the thin metal film. In this case, the number of waves of the surface plasmon resonance depends on the dielectric ratio of the thin metal film and a refraction factor of a sample fixed to be in contact with a surface of the prism having the thin metal film. Thus, it is possible to check the refraction factor and a dielectric ratio of the sample. That is, when the optical system and the sample are located at opposite positions sandwiching the thin metal film, it is easy to constitute a sensor.
In connection with the aforementioned principle, there has been developed an SPR sensor for an immunoassay apparatus using an optical fiber (trade name: BIACORE Probe produced by the BIACORE CO., Ltd.). This SPR sensor using an optical fiber is produced as follows. Firstly, a clad is removed from the end of the optical fiber and the optical fiber end surface, i.e., the core end surface is accurately cut or polished and this end surface is coated with silver. Moreover, the exposed core portion after removing the clad is covered with a thin metal film (gold or silver). Furthermore, he thin metal film is covered with a dielectric film, onto which an antibody used for the immunoassay is fixed. Moreover, at the other end of the optical fiber, a predetermined light source is provided so as to introduce a light into the optical fiber.
Description will now be directed to the immunoassay technique using the SPR sensor having the aforementioned configuration.
Firstly, the light from the light source is introduced into the optical fiber and a light of specific wavelength excites a surface plasmon resonance at the end of the optical fiber. The wavelength causing the surface plasmon resonance is changed by the refraction factor between the dielectric film and the antigen. The light of the wavelength which has caused the surface plasmon resonance is attenuated. Thus, the immunoassay can be performed by comparing the light wavelength attenuated most prior to the immune reaction and the light wavelength attenuated most after the immune reaction.
In addition to the SPR sensor using the optical fiber, there has been developed an SPR sensor using a prism.
However, the aforementioned conventional sensors have various problems and disadvantages. When an SPR sensor is constituted by using an optical fiber, it is necessary to cover the optical fiber core end portion with a thin metal film (for example, Au deposition). However, the optical fiber itself is a very small component and it is not easy to form a thin metal film appropriately.
Moreover, when performing an immunoassay actually, an antibody should be fixed onto the surface of the thin metal film. However, the optical fiber core has a cylindrical shape and it is not easy to fix the antibody.
Moreover, in the immunoassay apparatus using a conventional SPR sensor, there is provided only one SPR sensor having a single optical fiber. This brings about various problems. When using the enzyme immunoassay (EIA), a number of measurement steps are required and a long time is required for the immune reaction. That is, a measurement of one sample requires several hours or several tens of hours, and it is impossible to increase the measurement efficiency.
Moreover, in the immunoassay apparatus using the SPR sensor having an optical fiber, an antibody to be used for an immune reaction is fixed to the end portion of the optical fiber so that an antigen to be assayed in the sample is subjected to a reaction with the antibody of the SPR sensor. That is, if a sample contains a number of antigens to be assayed, the antigens should be assayed successively one after another. Moreover, in the conventional immunoassay apparatus, when changing the measurement item, the entire optical fiber is should be replaced with another optical fiber.
SUMMARY OF THE INVENTION
It is therefore an object of the present invention to provide an SPR (surface plasmon resonance) sensor cell enabling to easily fix and maintain an antibody. The present invention also provides an immunoassay apparatus capable of performing an immunoassay quickly.
The SPR sensor cell according to the present invention comprises: a core transparent for a light; a clad covering the core and having a through hole at a predetermined position to communicate with the core; and a predetermined thin metal film formed on an exposed surface of the core corresponding to the through hole.
When performing an immunoassay, firstly, an antibody is fixed via a dielectric film on the thin metal film formed on the SPR sensor cell. Next, light is introduced into the SPR sensor cell so as to identify a wavelength of the light whose intensity is attenuated. After this, a the through hole is filled with a sample for immunoassay so as to obtain an immune reaction. Again, light is introduced into the SPR sensor cell so as to identify a wavelength of the light which is attenuated
Nakamura Muneaki
Negami Mitsuhiro
Suzuki Kazuyasu
McGinn & Gibb PLLC
Sines Brian
Suzuki Motor Corporation
Warden Jill
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