JP2000028608A - Kit for periodontitis diagnosis - Google Patents
Kit for periodontitis diagnosisInfo
- Publication number
- JP2000028608A JP2000028608A JP18188199A JP18188199A JP2000028608A JP 2000028608 A JP2000028608 A JP 2000028608A JP 18188199 A JP18188199 A JP 18188199A JP 18188199 A JP18188199 A JP 18188199A JP 2000028608 A JP2000028608 A JP 2000028608A
- Authority
- JP
- Japan
- Prior art keywords
- periodontal pocket
- tris
- periodontal
- hydrochloric acid
- buffer solution
- 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
Abstract
Description
【０００１】[0001]
【発明の属する技術分野】この発明は、歯周病の診断キ
ットに関し、特に血液中蛋白を高感度で測定することに
より歯周病の診断を行うキットに関する。The present invention relates to a kit for diagnosing periodontal disease, and more particularly to a kit for diagnosing periodontal disease by measuring blood protein with high sensitivity.
【０００２】[0002]
【従来の技術】歯周病は治癒可能な時期に早期発見して
治療を開始することが歯を失わないために重要である。
歯周病の診断は従来、歯周ポケットの深さを用手法で測
定する方法や歯周ポケットに存在する歯周病関連病原菌
を検出する方法が主に行われてきている。2. Description of the Related Art It is important to detect periodontal disease at an early stage when it can be cured and to start treatment, in order not to lose teeth.
Conventionally, diagnosis of periodontal disease has mainly been performed by a method of measuring the depth of a periodontal pocket by a manual method or a method of detecting periodontal disease-related pathogenic bacteria present in the periodontal pocket.
【０００３】[0003]
【発明が解決しようとする課題】しかしながら、歯周ポ
ケットの深さを用手法で測定する方法で精度の良い測定
をするためには測定する人間の熟練が必要であり、人手
に頼るため、処理能力が低いという欠点がある。また、
歯周ポケットの歯周病関連病原菌を検出する方法は大量
処理が可能であるが、原因となる菌が１種類ではないこ
となどの理由ですべての歯周病を診断することは困難で
ある。この発明は、これらの問題点に着目してなされた
ものであって、上記の従来の方法にかわる歯周病の診断
キットを提供することを目的とする。However, in order to measure the depth of the periodontal pocket by using the method with high accuracy, it is necessary to have a skill of the person who performs the measurement. There is a drawback of low ability. Also,
Although the method for detecting periodontal disease-related pathogenic bacteria in the periodontal pocket can be processed in a large amount, it is difficult to diagnose all periodontal diseases because the causative bacteria are not one kind. The present invention has been made in view of these problems, and an object of the present invention is to provide a diagnostic kit for periodontal disease which can be replaced by the above-mentioned conventional method.
【０００４】[0004]
【課題を解決するための手段】上記の目的を達成するた
めに、この発明では、歯周組織からの出血により歯周ポ
ケット中に増加する血液中蛋白を測定対象としている。
微量の出血を検出するためにはアルファ１−アンチトリ
プシンあるいはアルブミンなどの血液中の濃度の高い蛋
白が測定対象として適当であり、アルファ１−アンチト
リプシンは蛋白分解酵素の影響を受けにくいことから、
もっとも適当な測定対象となりうる。なお歯周ポケット
中のアルファ１−アンチトリプシンなどの血液中蛋白を
高感度で測定するには、酵素免疫法やラテックス凝集反
応、免疫比濁法など日常、多用されている免疫学的測定
法を用いることができる。測定対象の試料としては歯周
滲出液、歯周ポケット部プラーク、唾液などの歯周ポケ
ット成分が含有される試料が利用できる。In order to achieve the above object, in the present invention, a protein in blood which increases in a periodontal pocket due to bleeding from periodontal tissue is measured.
In order to detect a small amount of bleeding, a protein having a high concentration in the blood such as alpha 1-antitrypsin or albumin is suitable for measurement, and alpha 1-antitrypsin is less susceptible to proteolytic enzymes.
It can be the most appropriate measurement target. In addition, in order to measure blood proteins such as alpha 1-antitrypsin in periodontal pockets with high sensitivity, immunological measurement methods that are commonly used in daily life, such as enzyme immunoassay, latex agglutination, and immunoturbidimetry, are used. Can be used. As a sample to be measured, a sample containing periodontal pocket components such as periodontal exudate, periodontal pocket plaque, and saliva can be used.
【０００５】今回、本発明者は、歯周病において歯周ポ
ケット中の血清中蛋白が正常よりも高値を示し、なおか
つ病状の悪化で増加することを発見した（図１）。ま
た、歯周ポケットの深さを中心に歯周病診断を行うＣＰ
ＩＴＮ法とも関連性が高いことが確認された（図２）。
以上のごとく、歯周ポケット中の血液中蛋白を高感度で
測定することにより、歯周病の診断ができる。[0005] The present inventors have now found that in periodontal disease, the serum protein in the periodontal pocket shows a higher level than normal, and increases due to worsening of the disease state (Fig. 1). In addition, a CP for performing periodontal disease diagnosis centering on the depth of the periodontal pocket
It was confirmed that it is highly related to the ITN method (FIG. 2).
As described above, periodontal disease can be diagnosed by measuring the blood protein in the periodontal pocket with high sensitivity.
【０００６】[0006]
【実施例】酵素免疫法によるアルファ１−アンチトリプ
シンの測定 〔マイクロプレートへの抗体の固相化〕マイクロプレー
トの各ウェルに抗アルファ１−アンチトリプシン抗体５
μg/mlを含むトリス−塩酸緩衝液（0.05M pH 8.4）を 1
00μl ずつ分注し、一夜４℃で放置して抗体を物理吸着
の原理でウェルに固相化する。 〔酵素標識抗体の調製〕別途、過ヨーソ酸法により、ア
ルカリホスファターゼを抗アルファ１−アンチトリプシ
ン抗体に標識して調製する。 〔歯周ポケット試料の採取〕細い濾紙片を歯周ポケット
に差し込み、歯周ポケット成分を染み込ませ、容器中で
0.1％の牛血清アルブミンを含むトリス−塩酸緩衝液
（0.05M pH 7.5）を用いてアルファ１−アンチトリプシ
ンを抽出する。[Example] Measurement of alpha 1-antitrypsin by enzyme immunoassay [Immobilization of antibody on microplate] Anti-alpha1-antitrypsin antibody 5 was added to each well of the microplate.
1 μl / ml Tris-HCl buffer (0.05M pH 8.4)
Dispense 00 μl each and leave it overnight at 4 ° C. to immobilize the antibody on the wells by the principle of physical adsorption. [Preparation of Enzyme-Labeled Antibody] Separately, anti-alpha 1-antitrypsin antibody is labeled with alkaline phosphatase and prepared by the periodate method. [Sampling of periodontal pocket sample] Insert a small piece of filter paper into the periodontal pocket, soak the periodontal pocket component,
Alpha 1-antitrypsin is extracted using Tris-HCl buffer (0.05 M pH 7.5) containing 0.1% bovine serum albumin.
【０００７】〔歯周ポケット中ラクトフェリンの測定〕
各ウェルに歯周ポケット成分含有サンプルを 100μl 分
注し、混和した後、37℃で１時間反応させる。次にマイ
クロプレートをTween 20を0.05％含むトリス−塩酸緩衝
液（0.05M pH7.5）で３回洗浄する。その後、酵素標識
抗アルファ１−アンチトリプシン抗体2.5μg/mlおよび
１％の牛血清アルブミンを含むトリス−塩酸緩衝生理食
塩水（0.1M pH 8.0)を 100μl ずつ分注し、混和した
後、37℃で１時間反応させる。次にマイクロプレートを
Tween 20を0.05％含むトリス−塩酸緩衝液（0.05M pH7.
5）で３回洗浄する。その後、Kind-King 法の基質緩衝
液 100μl を各ウェルに加え、37℃で30分間反応させ
る。ここで基質緩衝液は、Disodium Phenyl phosphate
0.215gと4-aminoantipyrine 0.09g を炭酸緩衝液（0.05
M pH 10.15）100ml に溶解したものである。次いで、 1
00μl の呈色液を各ウェルに加えて、呈色させる。ここ
で呈色液は、200ml の脱イオン水に2.6gのホウ酸を溶解
させた後、0.38g のPotassium Ferricyanideを溶解させ
たものである。最後に、各ウェルの呈色をマイクロプレ
ート用比色計を用いて510/630nm の波長で比色し検量線
から歯周ポケット中アルファ１−アンチトリプシン濃度
を算出する。[Measurement of lactoferrin in periodontal pocket]
100 μl of a sample containing periodontal pocket components is dispensed into each well, mixed, and reacted at 37 ° C. for 1 hour. Next, the microplate is washed three times with a Tris-HCl buffer (0.05 M pH 7.5) containing 0.05% Tween 20. Thereafter, 100 μl of Tris-HCl buffered saline (0.1 M pH 8.0) containing 2.5 μg / ml of enzyme-labeled anti-alpha 1-antitrypsin antibody and 1% bovine serum albumin was dispensed and mixed, and then mixed at 37 ° C. And react for 1 hour. Next, the microplate
Tris-HCl buffer containing 0.05% Tween 20 (0.05M pH7.
Wash 3 times in 5). Thereafter, 100 μl of a substrate buffer of the Kind-King method is added to each well and reacted at 37 ° C. for 30 minutes. Here, the substrate buffer is Disodium Phenyl phosphate
0.215 g and 0.09 g of 4-aminoantipyrine were added to a carbonate buffer (0.05
M pH 10.15) Dissolved in 100 ml. Then 1
Add 00 μl of color solution to each well to allow color development. Here, the color solution is obtained by dissolving 2.6 g of boric acid in 200 ml of deionized water and then dissolving 0.38 g of Potassium ferricyanide. Finally, the color of each well is compared with a colorimeter for microplate at a wavelength of 510/630 nm, and the concentration of alpha 1-antitrypsin in the periodontal pocket is calculated from the calibration curve.
【０００８】[0008]
【発明の効果】以上説明したように、歯周ポケット中の
血液中蛋白を高感度で測定すれば、歯周病の診断および
病状の把握ができる。As described above, by measuring the protein in the blood in the periodontal pocket with high sensitivity, it is possible to diagnose periodontal disease and to grasp its condition.
【図１】歯周ポケット中の血清中蛋白値の比較図であ
る。FIG. 1 is a graph comparing serum protein levels in periodontal pockets.
【図２】歯周ポケット中の血清中蛋白値とＣＰＩＴＮ法
の関連性を示す図面である。FIG. 2 is a drawing showing the relationship between the serum protein level in the periodontal pocket and the CPITN method.
Claims (1)
チトリプシン抗体などの抗体を備えて、歯周滲出液中に
おける血液蛋白を測定対象として、酵素免疫法、ラテッ
クス凝集反応、免疫比濁法などの免疫学的方法を用いる
歯周病の診断キット。1. An antibody, such as an anti-albumin antibody or an anti-alpha 1-antitrypsin antibody, which is used for measuring blood proteins in periodontal exudate, such as enzyme immunoassay, latex agglutination, and immunoturbidimetry. A diagnostic kit for periodontal disease using an immunological method.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP18188199A JP2000028608A (en) | 1999-06-28 | 1999-06-28 | Kit for periodontitis diagnosis |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP18188199A JP2000028608A (en) | 1999-06-28 | 1999-06-28 | Kit for periodontitis diagnosis |
Related Parent Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP01450597A Division JP3165789B2 (en) | 1997-01-09 | 1997-01-09 | Periodontal disease diagnostic kit |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| JP2000028608A true JP2000028608A (en) | 2000-01-28 |
Family
ID=16108513
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP18188199A Pending JP2000028608A (en) | 1999-06-28 | 1999-06-28 | Kit for periodontitis diagnosis |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JP2000028608A (en) |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2003066039A (en) * | 2001-08-22 | 2003-03-05 | Univ Nihon | Examination method of periodontitis by quantitative determination of liver cell growth factor in sputum |
| WO2010122946A1 (en) | 2009-04-24 | 2010-10-28 | 花王株式会社 | Periodontal disease markers |
| CN102998448A (en) * | 2011-09-09 | 2013-03-27 | 上海市计划生育科学研究所 | Prostatic cancer markers from prostatic secretion |
| CN104374924A (en) * | 2014-12-15 | 2015-02-25 | 山东博科生物产业有限公司 | Alpha1-AT (antitrypsin) immunoturbidimetry detection kit |
-
1999
- 1999-06-28 JP JP18188199A patent/JP2000028608A/en active Pending
Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2003066039A (en) * | 2001-08-22 | 2003-03-05 | Univ Nihon | Examination method of periodontitis by quantitative determination of liver cell growth factor in sputum |
| JP4696252B2 (en) * | 2001-08-22 | 2011-06-08 | 学校法人日本大学 | Examination method of periodontitis by quantification of hepatocyte growth factor in saliva |
| WO2010122946A1 (en) | 2009-04-24 | 2010-10-28 | 花王株式会社 | Periodontal disease markers |
| US8293478B2 (en) | 2009-04-24 | 2012-10-23 | Kao Corporation | Method of using autoinducer-2 as a periodontal disease marker |
| EP2631648A2 (en) | 2009-04-24 | 2013-08-28 | Kao Corporation | Therapeutic target for periodontal disease. |
| US8568986B2 (en) | 2009-04-24 | 2013-10-29 | Kao Corporation | Method of selecting an agent that improves periodontal disease |
| CN102998448A (en) * | 2011-09-09 | 2013-03-27 | 上海市计划生育科学研究所 | Prostatic cancer markers from prostatic secretion |
| CN104374924A (en) * | 2014-12-15 | 2015-02-25 | 山东博科生物产业有限公司 | Alpha1-AT (antitrypsin) immunoturbidimetry detection kit |
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Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| A02 | Decision of refusal |
Free format text: JAPANESE INTERMEDIATE CODE: A02 Effective date: 20011211 |