JP3036063B2 - How to measure fibrin gel dissolution time - Google Patents
How to measure fibrin gel dissolution timeInfo
- Publication number
- JP3036063B2 JP3036063B2 JP2316290A JP31629090A JP3036063B2 JP 3036063 B2 JP3036063 B2 JP 3036063B2 JP 2316290 A JP2316290 A JP 2316290A JP 31629090 A JP31629090 A JP 31629090A JP 3036063 B2 JP3036063 B2 JP 3036063B2
- Authority
- JP
- Japan
- Prior art keywords
- fibrin gel
- time
- absorbance
- light
- reaction
- 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.)
- Expired - Lifetime
Links
- 102000009123 Fibrin Human genes 0.000 title claims description 45
- 108010073385 Fibrin Proteins 0.000 title claims description 45
- BWGVNKXGVNDBDI-UHFFFAOYSA-N Fibrin monomer Chemical compound CNC(=O)CNC(=O)CN BWGVNKXGVNDBDI-UHFFFAOYSA-N 0.000 title claims description 45
- 229950003499 fibrin Drugs 0.000 title claims description 45
- 238000004090 dissolution Methods 0.000 title claims description 24
- 238000002835 absorbance Methods 0.000 claims description 18
- 238000002834 transmittance Methods 0.000 claims description 17
- 108010001014 Plasminogen Activators Proteins 0.000 claims description 12
- 102000001938 Plasminogen Activators Human genes 0.000 claims description 12
- 239000011324 bead Substances 0.000 claims description 12
- 238000000034 method Methods 0.000 claims description 12
- 229940127126 plasminogen activator Drugs 0.000 claims description 12
- 239000011521 glass Substances 0.000 claims description 9
- 102000013566 Plasminogen Human genes 0.000 claims description 8
- 108010051456 Plasminogen Proteins 0.000 claims description 8
- 108010049003 Fibrinogen Proteins 0.000 claims description 7
- 102000008946 Fibrinogen Human genes 0.000 claims description 7
- 108090000190 Thrombin Proteins 0.000 claims description 7
- 229940012952 fibrinogen Drugs 0.000 claims description 7
- 239000000126 substance Substances 0.000 claims description 7
- 229960004072 thrombin Drugs 0.000 claims description 7
- 230000005484 gravity Effects 0.000 claims description 4
- 230000000694 effects Effects 0.000 description 13
- 239000000243 solution Substances 0.000 description 12
- 238000005259 measurement Methods 0.000 description 11
- 102000004190 Enzymes Human genes 0.000 description 6
- 108090000790 Enzymes Proteins 0.000 description 6
- 229940088598 enzyme Drugs 0.000 description 6
- 230000005540 biological transmission Effects 0.000 description 3
- 238000000691 measurement method Methods 0.000 description 3
- 208000007536 Thrombosis Diseases 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 230000015271 coagulation Effects 0.000 description 2
- 238000005345 coagulation Methods 0.000 description 2
- 230000023597 hemostasis Effects 0.000 description 2
- 206010002383 Angina Pectoris Diseases 0.000 description 1
- 206010008132 Cerebral thrombosis Diseases 0.000 description 1
- 201000001429 Intracranial Thrombosis Diseases 0.000 description 1
- 108010023197 Streptokinase Proteins 0.000 description 1
- 108090000373 Tissue Plasminogen Activator Proteins 0.000 description 1
- 102000003978 Tissue Plasminogen Activator Human genes 0.000 description 1
- 108090000435 Urokinase-type plasminogen activator Proteins 0.000 description 1
- 102000003990 Urokinase-type plasminogen activator Human genes 0.000 description 1
- 125000003275 alpha amino acid group Chemical group 0.000 description 1
- -1 as shown below Proteins 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 210000000601 blood cell Anatomy 0.000 description 1
- 238000004113 cell culture Methods 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 230000009089 cytolysis Effects 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 238000001879 gelation Methods 0.000 description 1
- 230000001678 irradiating effect Effects 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000002844 melting Methods 0.000 description 1
- 230000008018 melting Effects 0.000 description 1
- 239000002184 metal Substances 0.000 description 1
- 239000011259 mixed solution Substances 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 229940012957 plasmin Drugs 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 238000004445 quantitative analysis Methods 0.000 description 1
- 230000006798 recombination Effects 0.000 description 1
- 229960005202 streptokinase Drugs 0.000 description 1
- 238000010998 test method Methods 0.000 description 1
- 229940124597 therapeutic agent Drugs 0.000 description 1
- 229960000187 tissue plasminogen activator Drugs 0.000 description 1
- 229960005356 urokinase Drugs 0.000 description 1
Landscapes
- Investigating Or Analysing Biological Materials (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Description
【発明の詳細な説明】 ［産業上の利用分野］ この発明はフィブリンゲル溶解時間の測定方法に関す
るものであり、医療などの分野で利用される。The present invention relates to a method for measuring a fibrin gel dissolution time, and is used in fields such as medical treatment.
［従来技術］ 脳血栓や狭心症治療薬としてヒト組織プラスミノーゲ
ン活性化因子（以下tPAという）が開発されている。こ
のtPAは各メーカーにより遺伝子組替えや細胞培養など
それぞれ異なった方法で製造しているため、製造された
tPAのアミノ酸配列や分子量が異なり、効果が微妙に違
っている。そのため、このtPAの効力を同じ試験法によ
って調べることにより、効力の判定を統一する必要があ
る。[Prior Art] Human tissue plasminogen activator (hereinafter referred to as tPA) has been developed as a therapeutic agent for cerebral thrombosis and angina. This tPA is manufactured by each manufacturer in different ways such as genetic recombination and cell culture,
The amino acid sequence and molecular weight of tPA are different, and the effects are slightly different. Therefore, it is necessary to unify the determination of efficacy by examining the efficacy of this tPA by the same test method.
従来tPAなどのプラスミノーゲンアクチベーターの活
性測定法として、クロットリシスタイム法が用いられて
いる。この方法は以下に示すようにフィブリノーゲン、
トロンビン、プラスミノーゲン及びプラスミノーゲンア
クチベーターの４種のタンパク質を混合し、これらのう
ちフィブリノーゲンとトロンビンの反応によりフィブリ
ンゲルを生成させ、その後プラスミノーゲンとプラスミ
ノーゲンアクチベーターの反応により生じたプラスミン
によりフィブリンゲルが溶解する現象を利用した定量法
である。Conventionally, a clot lysis time method has been used as a method for measuring the activity of a plasminogen activator such as tPA. This method uses fibrinogen, as shown below,
Thrombin, plasminogen, and plasminogen activator are mixed, and the fibrin gel is formed by the reaction of fibrinogen and thrombin, and then generated by the reaction of plasminogen and plasminogen activator. This is a quantitative method using the phenomenon that fibrin gel is dissolved by plasmin.
この測定は、試験管内に調整したフィブリンゲル上に
ガラスビーズを載せておくとゲルが溶解した時点でこの
ビーズが落下する。この落下までの時間がプラスミノー
ゲンアクチベーターの活性に依存するので、上記４種の
タンパク質混合時をスタートとして、ビーズ落下時まで
の時間を測定者が目視により、ストップウォッチを用い
て測定することにより活性測定を行なっていた。 In this measurement, when glass beads are placed on a fibrin gel prepared in a test tube, the beads fall when the gel is dissolved. Since the time until this drop depends on the activity of the plasminogen activator, the time until the beads fall from the start of mixing the above four proteins should be measured visually by a measurer using a stopwatch. Was used to measure the activity.
［発明が解決しようとする課題］ 従来方法では、測定者が目視によりビーズ落下までの
時間を測定してため、測定者が替わることによる測定時
間のバラツキがあり、また同一測定者であっても、長時
間の測定からくる疲労に伴う測定時間のバラツキがあっ
た。[Problem to be Solved by the Invention] In the conventional method, since the measurer measures the time until the beads fall visually, there is a variation in the measurement time due to the change of the measurer, and even if the same measurer is used. In addition, there was variation in measurement time due to fatigue caused by long-time measurement.
さらに、一度に多くのサンプルを測定する場合には、
測定者がこの測定に拘束されて、別の作業ができないと
いう問題点もあった。Furthermore, when measuring many samples at once,
There is also a problem that the measurer is restricted by this measurement and cannot perform another work.
［課題を解決するための手段］ この発明は上記の点に鑑みてなされたものであり、フ
ィブリンゲルの溶解時間を光学的に自動測定することに
より、プラスミノーゲンアクチベーター活性を求めるこ
とを目的としたものである。Means for Solving the Problems The present invention has been made in view of the above points, and has as its object to determine the plasminogen activator activity by optically automatically measuring the dissolution time of fibrin gel. It is what it was.
この発明のフィブリンゲル溶解時間の測定方法は、試
験管などの容器に、フィブリノーゲン、トロンビン、プ
ラスミノーゲン及びプラスミノーゲンアクチベーターを
加えてフィブリンゲルを形成した後、フィブリンゲル上
に、フィブリンゲルがプラスミノーゲン及びプラスミノ
ーゲンアクチベーターによって溶解した際に落下するよ
うな比重を有し、かつフィブリンゲルの溶液とは異なる
光の透過率または吸光度を有する指示物質を載せ、フィ
ブリンゲルの溶解に伴う指示物質の落下を、光源から当
該容器に光を照射して、当該容器を透過した光の透過率
または吸光度を測定することで検知し、反応開始時の透
過率または吸光度の初期値を基準として、反応開始後の
ある時点の透過光量比または吸光度比が、予め設定した
値以下である場合に、その時点をフィブリンゲルの溶解
時間とするものである。In the method for measuring the fibrin gel dissolution time of the present invention, a fibrin gel is formed by adding fibrinogen, thrombin, plasminogen and plasminogen activator to a container such as a test tube to form a fibrin gel. Plasminogen and an indicator that has a specific gravity such that it drops when dissolved by plasminogen activator and has a light transmittance or absorbance different from that of the solution of fibrin gel are placed, and accompanying the dissolution of fibrin gel The fall of the indicator substance is detected by irradiating the container with light from a light source and measuring the transmittance or absorbance of the light transmitted through the container, based on the initial value of the transmittance or absorbance at the start of the reaction. When the transmitted light amount ratio or the absorbance ratio at a certain point after the start of the reaction is equal to or less than a preset value, The point is to the dissolution time of the fibrin gel.
この発明のフィブリンゲル溶解時間の測定方法につい
て、第１図および第２図に基いて詳細に説明する。The method for measuring the fibrin gel dissolution time of the present invention will be described in detail with reference to FIGS. 1 and 2.
まず、試験管（１）に、フィブリノーゲン、トロンビ
ンおよびプラスミノーゲン、プラスミノーゲンアクチベ
ーターをほぼ同時に加えて撹拌し、フィブリンゲル
（２）を生成させる。First, fibrinogen, thrombin, plasminogen, and plasminogen activator are almost simultaneously added to a test tube (1) and stirred to generate a fibrin gel (2).
次いで、フィブリンゲル（２）の上に、フィブリンゲ
ル（２）が溶解した際に落下するような比重を有し、か
つフィブリンゲル（２）の溶液とは異なる光の透過率ま
たは吸光度を有する指示物質（３）を載せる。Then, an indicator having a specific gravity on the fibrin gel (2) such that the fibrin gel (2) falls when dissolved, and having a different light transmittance or absorbance from the solution of the fibrin gel (2) Place substance (3).
次いで光源（４）から試験管（１）の底よりやや高い
位置に光を一定時間毎に照射し、試験管（１）を透過し
た光を光学的に測定、すなわち光の透過率または吸光度
を分光光度計（５）により測定する。第１図に示すよう
にフィブリンゲル（２）が溶解しない間は、指示物質
（３）は落下しないので光の透過率または吸光度は変化
しないが、第２図に示すように、フィブリンゲル（２）
が溶解して溶液（２′）になると指示物質（３）が落下
し、指示物質（３）に光が透過するので、透過率または
吸光度が変化する。Next, light is emitted from the light source (4) to a position slightly higher than the bottom of the test tube (1) at regular intervals, and the light transmitted through the test tube (1) is optically measured, that is, the transmittance or absorbance of the light is measured. It is measured by a spectrophotometer (5). While the fibrin gel (2) does not dissolve as shown in FIG. 1, the indicator (3) does not fall, so that the light transmittance or absorbance does not change. However, as shown in FIG. )
Is dissolved into a solution (2 '), the indicator (3) falls, and light passes through the indicator (3), so that the transmittance or the absorbance changes.
この明細書において、フィブリンゲルの溶液とは、フ
ィブリンゲルが完全に溶けた状態だけでなく、フィブリ
ンゲルが一部溶けていない状態も含むものとする。In this specification, the term “fibrin gel solution” includes not only a state in which fibrin gel is completely dissolved but also a state in which fibrin gel is not partially dissolved.
この発明のフィブリンゲル溶解時間の測定方法はこの
光の透過率または吸光度の変化に着目したものであり、
上記のフィブリンゲルを形成する試薬およびその凝固物
を溶解させる試薬を加えて撹拌した時点を反応開始時間
とし、フィブリンゲル生成後であって指示物質をフィブ
リンゲルの上に載せた時点での光の透過率または吸光度
の初期値を基準とし、反応開始後のある時点での透過光
量比［（反応開始後のある時点での透過率の値／透過率
の初期値）×100％］、または吸光度比［（反応開始後
のある時点での吸光度の値／吸光度の初期値）×100
％］が予め設定した値以下である場合に、その時点を凝
固物溶解時間とするものである。The method for measuring the fibrin gel dissolution time of the present invention focuses on this change in light transmittance or absorbance,
The time when the above-described reagent for forming the fibrin gel and the reagent for dissolving the coagulated substance were added and stirred was taken as the reaction start time, and the light emission at the time when the indicator was placed on the fibrin gel after fibrin gel formation was performed. Based on the initial value of the transmittance or the absorbance, the ratio of the amount of transmitted light at a certain point after the start of the reaction [(the value of the transmittance at a certain point after the start of the reaction / the initial value of the transmittance) × 100%], or the absorbance Ratio [(value of absorbance at a certain time after the start of reaction / initial value of absorbance) × 100
%] Is equal to or less than a preset value, the time is taken as the coagulation dissolution time.
プラスミノーゲンアクチベーターとしては、tPA、ウ
ロキナーゼ、ストレプトキナーゼなどが挙げられる。Plasminogen activators include tPA, urokinase, streptokinase and the like.
すなわちこの発明の測定方法によってフィブリンゲル
の溶解時間を測定することにより、上記プラスミノーゲ
ンアクチベーターの活性を測定することができる。That is, the activity of the plasminogen activator can be measured by measuring the dissolution time of the fibrin gel by the measurement method of the present invention.
「指示物質」としては、凝固物が溶解した際に落下す
るような比重を有し、かつ凝固物の溶液とは異なる光の
透過率または吸光度を有するものであれば特に限定され
ないが、例えば透明または着色されたガラスビーズ、金
属性の球などが挙げられる。The “indicator” is not particularly limited as long as it has a specific gravity such that it falls when the coagulate is dissolved, and has a light transmittance or absorbance different from that of the coagulate solution. Alternatively, colored glass beads, metal spheres, and the like can be given.
光源から照射される光の波長は、凝固物の色調や濃
度、指示物質の色調や濃度などにより任意に選択され
る。The wavelength of the light emitted from the light source is arbitrarily selected depending on the color tone and concentration of the coagulated material, the color tone and concentration of the indicator, and the like.
この凝固物溶解時間の測定は、例えば市販されている
トキシノメーターET−201（和光純薬工業株式会社製）
などを用いて行なうことができる。このトキシノメータ
ーET−201は、内部を一定温度に保つことのできる複数
の反応試験管セット用ホール、マイクロコンピュータ
ー、それぞれの試験管に対応する光源、分光光度計など
を有し、エンドトキシンとカブトガニの血球抽出成分と
のゲル化反応に伴う濁度変化を、一定波長における透過
光量の変化として捉えて測定する装置である。The measurement of the coagulation dissolution time is performed, for example, by using a commercially available toxinometer ET-201 (manufactured by Wako Pure Chemical Industries, Ltd.).
It can be performed using, for example. The toxinometer ET-201 has a plurality of reaction test tube setting holes capable of keeping the inside at a constant temperature, a microcomputer, a light source corresponding to each test tube, a spectrophotometer, and the like. This is a device that measures and measures the change in turbidity caused by the gelation reaction with the blood cell extraction component as a change in the amount of transmitted light at a certain wavelength.
次にこの発明の測定方法の実施例として、tPAの酵素
比活性測定方法の一例について説明する。Next, as an example of the measurement method of the present invention, an example of a method for measuring the enzyme specific activity of tPA will be described.
［実施例］ 酵素比活性が既知のtPAの溶液（濃度:25μg/ml、酵素
比活性:4.7×105U/mg）およびこの溶液をそれぞれ1/2、
1/4、1/8に希釈した溶液を調整する。同様に、酵素比活
性を測定しようとするtPAの溶液（25μg/ml）およびこ
の溶液をそれぞれ1/2、1/4、1/8に希釈した溶液を調整
する。８本の試験管に上記のように調整したtPAをそれ
ぞれ100μ加え、各試験管にトロンビン（100U/ml）を
30μ加える。次にフィブリノーゲン（4mg/ml）および
プラスミノーゲン（0.2CU/ml）の混液を1ml加えて撹拌
した時点を反応開始時間として設定する。[Example] A solution of tPA having a known enzyme specific activity (concentration: 25 μg / ml, enzyme specific activity: 4.7 × 10 5 U / mg) and this solution were halved,
Prepare 1/4, 1/8 diluted solution. Similarly, a solution of tPA (25 μg / ml) whose enzyme specific activity is to be measured and a solution obtained by diluting the solution to 1/2, 1/4, and 1/8, respectively, are prepared. To each of the eight test tubes, 100 μl of the tPA adjusted as described above was added, and thrombin (100 U / ml) was added to each test tube.
Add 30μ. Next, 1 ml of a mixed solution of fibrinogen (4 mg / ml) and plasminogen (0.2 CU / ml) is added, and the time when the mixture is stirred is set as the reaction start time.
次いで各試験管をトキシノメーターET−210（和光純
薬工業株式会社製）の反応試験管セット用ホール（37℃
に保たれている）に入れる。反応開始後１〜２分で各試
験管にフィブリノーゲンとトロンビンの反応によって生
じるフィブリンゲルが形成されるので、一旦ホールから
試験管を取り出して、このフィブリンゲル上に透明のガ
ラスビーズ（直径約5mm）を載せて、再びホールに入れ
る。光源より光（波長:660nm）を照射し、分光光度計に
よって光の透過率の初期値を測定し、マイクロコンピュ
ーターにその値を記憶させる。予めゲル溶解時間判定の
ための透過光量比［（ある時間における透過率の値／透
過率の初期値）×100％）］を50％に設定する。Next, each test tube was set in a reaction test tube setting hole (37 ° C.) of a toxinometer ET-210 (manufactured by Wako Pure Chemical Industries, Ltd.).
). One to two minutes after the start of the reaction, fibrin gel generated by the reaction of fibrinogen and thrombin is formed in each test tube. The test tube is once removed from the hole, and transparent glass beads (about 5 mm in diameter) are placed on the fibrin gel. And put it in the hall again. Light (wavelength: 660 nm) is irradiated from a light source, an initial value of light transmittance is measured by a spectrophotometer, and the value is stored in a microcomputer. The transmission light amount ratio [(transmission value at a certain time / initial transmission value) × 100%)] for gel dissolution time determination is set to 50% in advance.
次いで光源より0.2分間隔で光を照射し、各時間にお
ける光の透過率を測定し、マイクロコンピューターによ
り、初期値との比（透過光量比）を求め、プリントアウ
トする。Next, light is emitted from the light source at intervals of 0.2 minutes, the light transmittance at each time is measured, the ratio to the initial value (transmitted light amount ratio) is obtained by a microcomputer, and printed out.
第３図にプリントアウトされたデータの一例を示し、
左の数字は反応開始時間を、右の数字は透過光量比を示
している。反応開始後24.0分から24.8分までの透過光量
比は100％またはそれに近い値であり、ガラスビーズは
落下しておらずゲルはまだ溶解していない。FIG. 3 shows an example of the printed data,
The numbers on the left indicate the reaction start time, and the numbers on the right indicate the transmitted light amount ratio. The ratio of the amount of transmitted light from 24.0 minutes to 24.8 minutes after the start of the reaction is 100% or a value close thereto, and the glass beads have not dropped and the gel has not yet dissolved.
反応開始後25.0分では透過光量比は66.28％であり、
ガラスビーズが落下し始めていることがわかるが、設定
値（透過光量比50％）より高い値であるので、下流溶解
時間とは判定されない。At 25.0 minutes after the start of the reaction, the transmitted light amount ratio was 66.28%,
Although it can be seen that the glass beads have begun to fall, since the value is higher than the set value (transmitted light amount ratio 50%), the downstream melting time is not determined.
反応開始後25.2分では透過光量比は9.71％であり、設
定値より低い値であるので、この時間をもってゲル溶解
時間と判定する。At 25.2 minutes after the start of the reaction, the transmitted light amount ratio is 9.71%, which is lower than the set value. Therefore, this time is determined as the gel dissolution time.
なお反応開始後25.2分から25.8分までは透過光量比が
増加し、26.0分以降一定値になっているが、これは反応
開始後25.2分ではガラスビーズの周囲の密度の濃い部分
に光が透過し、26.0分でガラスビーズが完全に試験管の
底に落下してガラスビーズの中央部の密度の薄い部分に
光が透過しているためと推定される。The ratio of the amount of transmitted light increased from 25.2 minutes to 25.8 minutes after the start of the reaction, and became constant after 26.0 minutes.However, at 25.2 minutes after the start of the reaction, light was transmitted through the dense part around the glass beads. It is presumed that the glass beads completely dropped to the bottom of the test tube in 26.0 minutes, and light was transmitted through the low-density portion at the center of the glass beads.
このようにして全てのサンプルにつて表２のようにフ
ィブリンゲルの溶解時間を求め、次いでThrombosis and
Haemostasis［スロンボシス アンド ヘモスタシス、
58（４）1085〜1087（1987）］に記載の統計分析法に従
って計算して、tPAの酵素比活性（4.9×105U/mg）を求
める。In this way, the dissolution time of the fibrin gel was determined for all the samples as shown in Table 2, and then the Thrombosis and
Haemostasis [thrombosis and hemostasis,
58 (4) 1085-1087 (1987)] to determine the specific enzyme activity of tPA (4.9 × 10 5 U / mg).
この実施例では、凝固物測定時間の判定およびその測
定時間からtPAの酵素比活性の計算をヒトが行なってい
るが、これらをマイクロコンピューターに予めプログラ
ムしておいて自動的に判定および計算ができるようにし
ても良い。 In this example, the determination of the coagulate measurement time and the calculation of the enzyme specific activity of tPA from the measurement time are performed by a human, but these can be automatically determined and calculated by previously programming them in a microcomputer. You may do it.
［効果］ この発明のフィブリンゲル溶解時間の測定方法によれ
ば、フィブリンゲルの溶解に伴う指示物質の落下を光学
的に自動測定できるので、測定者はこの測定に拘束され
ることなく別の作業ができる。また同一測定者における
測定時間のバラツキおよび測定者が異なることによる測
定時間のバラツキがないので、フィブリンゲル溶解時間
を正確に測定することができる。[Effect] According to the method for measuring the fibrin gel dissolution time of the present invention, the fall of the indicator substance accompanying the dissolution of the fibrin gel can be automatically measured optically, so that the measurer can perform another work without being restricted by this measurement. Can be. Further, since there is no variation in the measurement time of the same subject and no variation in the measurement time due to different subjects, the fibrin gel dissolution time can be accurately measured.
第１図および第２図はこの発明のフィブリンゲル溶解時
間の測定方法の概略説明図であり、第１図はフィブリン
ゲル溶解前、第２図はフィブリンゲル溶解後の状態をそ
れぞれ示す。第３図はこの測定方法における反応開始後
の時間と透過光量比を示すデータの一部を示す。 （１）試験管、（２）フィブリンゲル、（２′）フィブ
リンゲルの溶液 （３）指示物質、（４）光源、（５）分光光度計FIGS. 1 and 2 are schematic illustrations of the method for measuring the fibrin gel dissolution time of the present invention. FIG. 1 shows the state before fibrin gel dissolution, and FIG. 2 shows the state after fibrin gel dissolution. FIG. 3 shows a part of data indicating the ratio of the amount of transmitted light to the time after the start of the reaction in this measurement method. (1) test tube, (2) fibrin gel, (2 ') solution of fibrin gel (3) indicator, (4) light source, (5) spectrophotometer
Claims (2)
トロンビン、プラスミノーゲン及びプラスミノーゲンア
クチベーターを加えてフィブリンゲルを形成した後、フ
ィブリンゲル上に、フィブリンゲルがプラスミノーゲン
及びプラスミノーゲンアクチベーターによって溶解した
際に落下するような比重を有し、かつフィブリンゲルの
溶液とは異なる光の透過率または吸光度を有する指示物
質を載せ、フィブリンゲルの溶解に伴う指示物質の落下
を、光源から当該容器に光を照射して当該容器を透過し
た光の透過率または吸光度を測定することで検知し、反
応開始時の透過率または吸光度の初期値を基準として、
反応開始後のある時点の透過光量比または吸光度比が、
予め設定した値以下である場合にその時点をフィブリン
ゲルの溶解時間とすることを特徴とするフィブリンゲル
溶解時間の測定方法。(1) Fibrinogen, a container such as a test tube,
After forming a fibrin gel by adding thrombin, plasminogen and plasminogen activator, the specific gravity on the fibrin gel is such that the fibrin gel falls when dissolved by plasminogen and plasminogen activator. And, and put the indicator having a different light transmittance or absorbance from the solution of the fibrin gel, and the fall of the indicator due to the dissolution of the fibrin gel, the light was irradiated from the light source to the container and transmitted through the container Detected by measuring light transmittance or absorbance, based on the initial value of transmittance or absorbance at the start of the reaction,
At a certain time after the start of the reaction, the transmitted light amount ratio or the absorbance ratio is
A method for measuring a fibrin gel dissolution time, characterized in that when the value is equal to or less than a preset value, the time point is defined as a fibrin gel dissolution time.
（１）に記載のフィブリンゲル溶解時間の測定方法。2. The method for measuring a fibrin gel dissolution time according to claim 1, wherein the indicator substance is glass beads.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP2316290A JP3036063B2 (en) | 1990-11-20 | 1990-11-20 | How to measure fibrin gel dissolution time |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| JP2316290A JP3036063B2 (en) | 1990-11-20 | 1990-11-20 | How to measure fibrin gel dissolution time |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| JPH04187099A JPH04187099A (en) | 1992-07-03 |
| JP3036063B2 true JP3036063B2 (en) | 2000-04-24 |
Family
ID=18075465
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| JP2316290A Expired - Lifetime JP3036063B2 (en) | 1990-11-20 | 1990-11-20 | How to measure fibrin gel dissolution time |
Country Status (1)
| Country | Link |
|---|---|
| JP (1) | JP3036063B2 (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2015098473A1 (en) * | 2013-12-25 | 2015-07-02 | 株式会社日立ハイテクノロジーズ | Automatic analysis device and analysis method |
Families Citing this family (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| DE19608687A1 (en) * | 1996-03-06 | 1997-09-11 | Retro Tech Gmbh | Method and test kit for the non-radioactive, enzymatic detection of reverse transcriptase |
| US20220373566A1 (en) * | 2019-10-17 | 2022-11-24 | Nova Biomedical Corporation | Coagulation assay apparatus and methods thereof |
| JPWO2023074864A1 (en) * | 2021-10-29 | 2023-05-04 |
-
1990
- 1990-11-20 JP JP2316290A patent/JP3036063B2/en not_active Expired - Lifetime
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2015098473A1 (en) * | 2013-12-25 | 2015-07-02 | 株式会社日立ハイテクノロジーズ | Automatic analysis device and analysis method |
| JPWO2015098473A1 (en) * | 2013-12-25 | 2017-03-23 | 株式会社日立ハイテクノロジーズ | Automatic analyzer and analysis method |
| US10295555B2 (en) | 2013-12-25 | 2019-05-21 | Hitachi High-Technologies Corporation | Automatic analysis device and analysis method |
Also Published As
| Publication number | Publication date |
|---|---|
| JPH04187099A (en) | 1992-07-03 |
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