JPH04198865A - Immune measurement - Google Patents
Immune measurementInfo
- Publication number
- JPH04198865A JPH04198865A JP33222490A JP33222490A JPH04198865A JP H04198865 A JPH04198865 A JP H04198865A JP 33222490 A JP33222490 A JP 33222490A JP 33222490 A JP33222490 A JP 33222490A JP H04198865 A JPH04198865 A JP H04198865A
- Authority
- JP
- Japan
- Prior art keywords
- antibody
- ferritin
- hinokitiol
- reaction
- concentration
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 238000005259 measurement Methods 0.000 title description 6
- FUWUEFKEXZQKKA-UHFFFAOYSA-N beta-thujaplicin Chemical compound CC(C)C=1C=CC=C(O)C(=O)C=1 FUWUEFKEXZQKKA-UHFFFAOYSA-N 0.000 claims abstract description 22
- 102000008857 Ferritin Human genes 0.000 claims abstract description 16
- 108050000784 Ferritin Proteins 0.000 claims abstract description 16
- 238000008416 Ferritin Methods 0.000 claims abstract description 16
- TUFYVOCKVJOUIR-UHFFFAOYSA-N alpha-Thujaplicin Natural products CC(C)C=1C=CC=CC(=O)C=1O TUFYVOCKVJOUIR-UHFFFAOYSA-N 0.000 claims abstract description 11
- 229930007845 β-thujaplicin Natural products 0.000 claims abstract description 11
- 238000003018 immunoassay Methods 0.000 claims description 12
- 238000000034 method Methods 0.000 abstract description 15
- 239000012895 dilution Substances 0.000 abstract description 5
- 238000010790 dilution Methods 0.000 abstract description 5
- 239000007864 aqueous solution Substances 0.000 abstract description 4
- 102000004169 proteins and genes Human genes 0.000 abstract description 4
- 108090000623 proteins and genes Proteins 0.000 abstract description 4
- 239000000427 antigen Substances 0.000 abstract description 3
- 102000036639 antigens Human genes 0.000 abstract description 3
- 108091007433 antigens Proteins 0.000 abstract description 3
- 238000001514 detection method Methods 0.000 abstract description 2
- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 description 8
- 229910052742 iron Inorganic materials 0.000 description 4
- 238000011161 development Methods 0.000 description 3
- 102000004190 Enzymes Human genes 0.000 description 2
- 108090000790 Enzymes Proteins 0.000 description 2
- 210000004185 liver Anatomy 0.000 description 2
- 238000003127 radioimmunoassay Methods 0.000 description 2
- 230000035945 sensitivity Effects 0.000 description 2
- 102000000546 Apoferritins Human genes 0.000 description 1
- 108010002084 Apoferritins Proteins 0.000 description 1
- 210000001185 bone marrow Anatomy 0.000 description 1
- 239000004927 clay Substances 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000010894 electron beam technology Methods 0.000 description 1
- 210000003205 muscle Anatomy 0.000 description 1
- 238000001959 radiotherapy Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 239000000243 solution Substances 0.000 description 1
- 238000011179 visual inspection Methods 0.000 description 1
- 239000002699 waste material Substances 0.000 description 1
Abstract
Description
【発明の詳細な説明】 ［産業上の利用分野］ 本発明は免疫測定法に関する。[Detailed description of the invention] [Industrial application field] The present invention relates to immunoassays.
［従来の技術］
抗原又は抗体を検出、定量するための免疫測定法として
は、ラジオイムノアッセイ、エンザイムイムノアッセイ
、蛍光イムノアッセイ等があるが、なかでもエンザイム
イムノアツセイ（ＥＩＡ）はラジオイムノアッセイのよ
うに放射線の被爆や廃棄物処理等の問題がなく、大がか
りな設備を必要とせず、また、検出感度も満足できるも
のであるので広く普及している。しかしながら、ＥＩＡ
は測定者及び測定条件の変化により結果にばらつきが生
じるという欠点を有している。[Prior Art] Immunoassays for detecting and quantifying antigens or antibodies include radioimmunoassays, enzyme immunoassays, and fluorescence immunoassays, among which enzyme immunoassays (EIAs), like radioimmunoassays, do not require radiation therapy. It is widely used because there are no problems with radiation exposure or waste disposal, it does not require large-scale equipment, and its detection sensitivity is satisfactory. However, EIA
This method has the disadvantage that results vary due to changes in the measurer and measurement conditions.
一方、フェリチンを共有結合した抗体を用いた免疫測定
法も知られており、フェリチン抗体法とも呼ばれている
。フェリチンは肝臓、肝臓、骨髄及び筋肉組織に存在す
る分子量４６万の水溶性タンパク質であるアポフェリチ
ン１分子が２０００個の３価の鉄と結合したものである
。On the other hand, an immunoassay method using an antibody to which ferritin is covalently bound is also known, and is also called the ferritin antibody method. Ferritin is one molecule of apoferritin, which is a water-soluble protein with a molecular weight of 460,000 and is present in the liver, liver, bone marrow, and muscle tissues, bound to 2000 trivalent iron molecules.
フェリチンは多量に鉄を含むため、電子線を透過しにく
く、従って、電子顕微鏡で観察すると、フェリチンが存
在する部分は黒く見える。しかしながら、フェリチン抗
体法は、操作が煩雑で、電子顕微鏡のような高価で大が
かりな設備を必要とする。Since ferritin contains a large amount of iron, it is difficult for electron beams to pass through it, and therefore, when observed under an electron microscope, areas where ferritin exists appear black. However, the ferritin antibody method is complicated to operate and requires expensive and large-scale equipment such as an electron microscope.
［発明が解決しようとする問題点コ
従って、本発明の目的は、測定者や測定条件による結果
のばらつきがなく、特別な設備を必要とすることなく簡
単な操作で行なうことができる免疫測定法を提供するこ
とである。[Problems to be Solved by the Invention] Accordingly, an object of the present invention is to provide an immunoassay method that can be performed with simple operations without requiring any special equipment and without variations in results depending on the person performing the measurement or the measurement conditions. The goal is to provide the following.
［問題点を解決するための手段］
本願発明者らは、鋭意研究の結果、抗体と結合したフェ
リチン中の鉄分子とヒノキチオールが反応すると赤色に
発色し、この発色の程度を測定することにより免疫清１
定が可能であることを見出し本発明を完成した。[Means for Solving the Problems] As a result of intensive research, the inventors of the present application discovered that when hinokitiol reacts with iron molecules in ferritin bound to antibodies, a red color develops, and by measuring the degree of this color development, it was found that Sei 1
The present invention was completed based on the discovery that it is possible to
すなわち、本発明は、フェリチン標識した抗体をヒノキ
チオールと反応させて発色させる工程を含む免疫測定法
を提供する。That is, the present invention provides an immunoassay method that includes a step of reacting a ferritin-labeled antibody with hinokitiol to develop color.
［発明の詳細な説明］
本発明において用いられる、フェリチン標識した抗体は
、従来のフェリチン抗体法において用いられている方法
により作製することができ、土板のものを用いることも
できる。[Detailed Description of the Invention] The ferritin-labeled antibody used in the present invention can be produced by the method used in the conventional ferritin antibody method, and a clay plate antibody can also be used.
ヒノキチオールは、水溶液として抗体と反応させること
ができ、その濃度は、特に限定されないが例えば０，５
％程度である。Hinokitiol can be reacted with antibodies as an aqueous solution, and its concentration is not particularly limited, but for example, 0.5
It is about %.
フェリチン標識抗体と、ヒノキチオール水溶液との反応
は、室温で１分ないし１２０分程度で行なうことができ
る。The reaction between the ferritin-labeled antibody and the aqueous hinokitiol solution can be carried out at room temperature for about 1 to 120 minutes.
マイクロプレートを用いた免疫測定法の場合、目視可能
な発色を与えるためには、ヒノキチオールと反応する鉄
分子の数が約１０１５個以上必要である。従って、例え
ば、用いる抗体の蛋白濃度が６ ｍｇ／ｍｌの場合には
、＠出限界は約１５１１ｇ／ｍ１である。このように、
本発明の免疫−１１定法の感度は、従来の免疫測定法の
感度よりも高くはないが、試料を無希釈で用いることが
できるというｆｌ＋、占を有する。In the case of immunoassay using a microplate, the number of iron molecules that react with hinokitiol must be approximately 1015 or more in order to provide visible color development. Therefore, for example, when the protein concentration of the antibody used is 6 mg/ml, the output limit is about 1511 g/ml. in this way,
Although the sensitivity of the immuno-11 method of the present invention is not higher than that of conventional immunoassay methods, it has the advantage that the sample can be used without dilution.
本発明は、フェリチン標識した抗体をヒノキチオールと
の発色反応で検出する工程を含むあらゆる免疫測定法を
包含し、従来より行なわれている、直接法、間接法、サ
ンドイツチ法等のいずれの方法をも包含する。また、本
発明の方法により測定される抗原も何ら制限されるもの
ではない。The present invention encompasses all immunoassay methods that include the step of detecting ferritin-labeled antibodies through a color reaction with hinokitiol, and can be applied to any of the conventional methods such as the direct method, indirect method, and Sand-Deutsch method. include. Furthermore, the antigen that can be measured by the method of the present invention is not limited in any way.
以下、本発明を実施例によりさらに具体的に説明するが
、本発明は下記実施例に限定されるものではない。EXAMPLES Hereinafter, the present invention will be explained in more detail with reference to examples, but the present invention is not limited to the following examples.
し実施例］
重版のフェリチン標識抗ヒトＩｇＧ（蛋白濃度６ｉＩｇ
／ｍｌ 、ＥＹシラボラトリー社製）を２′倍に逓減希
釈したものをマイクロタイタープレートのウェルに入れ
、０．５％ヒノキチオール水溶液０．０２５ ｍｌを各
ウェルに入れ、室温で１分間反応させた後、目視にて発
色を＃！察した。Examples] Reprinted ferritin-labeled anti-human IgG (protein concentration 6iIg
/ml, manufactured by EY Syllatorial Co., Ltd.) was diluted 2' times and placed in the wells of a microtiter plate, and 0.025 ml of a 0.5% hinokitiol aqueous solution was placed in each well and reacted for 1 minute at room temperature. After that, check the color by visual inspection! I guessed it.
その結果、抗体濃度１５μｇ／ｍｌ （４００倍希釈）
以上の濃度で赤色の発色を観察することができた。As a result, the antibody concentration was 15μg/ml (400-fold dilution)
Red color development could be observed at the above concentrations.
［発明の効果］
本発明の方法によると、非常に簡便な操作で、測定者や
測定条件による結果のばらつきがなく、再現性良く抗体
を検出することができる。さらに、本発明の方法による
と、反応が迅速であるので、測定に要する時間が短い、
また、抗体試料を無希釈のまま用いることができるので
操作がより簡便である。[Effects of the Invention] According to the method of the present invention, antibodies can be detected with good reproducibility with very simple operations and without variations in results depending on the measurer or measurement conditions. Furthermore, according to the method of the present invention, since the reaction is rapid, the time required for measurement is short.
Furthermore, since the antibody sample can be used without dilution, the operation is simpler.
特許出願人 富士レビオ株式会社Patent applicant: Fujirebio Co., Ltd.
Claims (1)
発色させる工程を含む免疫測定法。An immunoassay method that includes the step of reacting a ferritin-labeled antibody with hinokitiol to develop color.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP33222490A JPH04198865A (en) | 1990-11-29 | 1990-11-29 | Immune measurement |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP33222490A JPH04198865A (en) | 1990-11-29 | 1990-11-29 | Immune measurement |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JPH04198865A true JPH04198865A (en) | 1992-07-20 |
Family
ID=18252563
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP33222490A Pending JPH04198865A (en) | 1990-11-29 | 1990-11-29 | Immune measurement |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JPH04198865A (en) |
-
1990
- 1990-11-29 JP JP33222490A patent/JPH04198865A/en active Pending
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