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Method of detecting platelet thrombosis or organ injury

A vascular and vascular technology, applied in the field of disseminated intravascular coagulation, can solve the problems of self-tissue damage, low specificity, tissue damage, etc., and achieve the effects of rapid detection, high specificity detection, and high clinical value.

Inactive Publication Date: 2007-10-17
MITSUBISHI KAGAKA IATRON INC
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, in tissues where an inflammatory response occurs, due to α 1 -AT is oxidized by reactive oxygen species released by neutrophils, myeloperoxidase, lactoferrin, which prevents neutrophil elastase from being inactivated, causing tissue damage
Due to the low substrate specificity of neutrophil elastase, when it is released in excess and α 1 - In the absence of inhibitors such as AT, there is a risk that the components of the organism will be decomposed, causing self-tissue damage caused by neutrophil elastase

Method used

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  • Method of detecting platelet thrombosis or organ injury
  • Method of detecting platelet thrombosis or organ injury
  • Method of detecting platelet thrombosis or organ injury

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] "Example 1: Decomposition of recombinant vWF decomposing enzyme antigen by elastase"

[0056] Dissolve 1.5 μg of recombinant vWF decomposing enzyme in tris buffer / physiological saline, add elastase until the final concentration is 20nmol / L or 140nmol / L, incubate at 37°C for 15 minutes or 1 hour, and decompose 0.5μg of vWF respectively enzyme. This was separated by non-reducing SDS electrophoresis (5 to 20% gel), and then transferred to a PVDF (polyvinylidene fluoride) membrane by the Western blot method. After blocking with a commercially available blocking agent (Block Ace, Dainippon Pharmaceutical Co., Ltd.) at room temperature for 30 minutes, they were washed with tris buffer. React in 1 μg / mL anti-vWF decomposing enzyme monoclonal antibody (WH2-22-1A: using disintegrin region of vWF decomposing enzyme as antigenic determinant) / tris buffer (pH7.4) / 10% Block Ace, react at room temperature After 1 hour, wash 3 times with tris buffer (pH7.4) / 0.05% NP-40, and then in 2...

Embodiment 2

[0065] "Example 2: Decomposition of plasmin or thrombin recombinant vWF decomposing enzyme antigen"

[0066] Dissolve 1.5 μg of recombinant vWF decomposing enzyme in tris buffer / normal saline, add plasminogen (final concentration 1 μmol / L) and tissue plasminogen activator (tPA) (final concentration 0.2nmol / L or 2nmol / L ), or add thrombin until the final concentration is 20mU or 200mU, and after incubating at 37°C for 15 minutes or 1 hour, take the equivalent of 0.5μg of vWF decomposing enzyme respectively. After it was separated by SDS electrophoresis, Western blot was performed in the same manner as in Example 1 to detect the band of vWF decomposing enzyme. The results are shown in Figure 2.

[0067] The corresponding relationship of each lane in Figure 2 is as follows:

[0068] 1: After adding 0.2nmol / L tPA, react at 37°C for 15 minutes

[0069] 2: After adding 2nmol / L tPA, react at 37°C for 15 minutes

[0070] 3: After adding 20mmol / L thrombin, react at 37°C for 15 minu...

Embodiment 3

[0078] "Example 3: Correlation between vWF decomposing enzyme and elastase"

[0079] In this embodiment, the plasma from normal healthy people, DIC patients and SIRS patients was used as samples to be tested, and the amount of vWF decomposing enzyme antigen and elastase were measured. The amount of vWF decomposing enzyme antigen was measured with a commercially available kit (vWF decomposing enzyme ELISA kit, Mitsubishi Chemical Yatron). In addition, the amount of elastase was expressed by measuring the amount of elastase / α1-antitrypsin using a commercially available kit (PMN Elastase / α1-PI Complex ELISA Kit, CALBIOCHEM).

[0080] The results are shown in Figures 3 and 4. In Fig. 3, the X-axis is the amount of elastase / α1-antitrypsin (unit: ng / mL), and the Y-axis is the amount of vWF decomposing enzyme antigen (unit: %). In addition, in Fig. 4, "N.S." indicates no significant difference.

[0081] A good negative correlation was shown between elastase / α1-antitrypsin and vWF ...

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Abstract

The present invention provides a method of detecting platelet thrombosis or organ injury in a patient suffering from disseminated intravascular coagulation (DIC) syndrome or systemic inflammatory response syndrome (SIRS) by analyzing a von Willebrand factor-degrading enzyme and / or a degrading factor thereof. A kit for detecting platelet thrombosis or organ injury in a patient suffering from DIC or SIRS which contains an antibody capable of binding specifically to a von Willebrand factor-degrading enzyme or its fragment and an antibody capable of binding specifically to a degrading factor of a von Willebrand factor-degrading enzyme or its fragment.

Description

technical field [0001] The present invention relates to a detection method of platelet thrombus or organ damage, especially the detection method for patients with disseminated intravascular coagulation (Disseminated Intravascular Coagulation; DIC) or systemic inflammatory response syndrome (Systemic inflammatory response syndrome; SIRS). According to the present invention, platelets can be detected by analyzing von Willebrand factor (vWF) and / or its decomposition factor in a biological sample (for example, blood) of a subject (in particular, a DIC or SIRS patient) Thrombosis or organ damage, for example, detecting or predicting the degree of platelet thrombus formation, predicting the incidence of thrombosis or organ damage (that is, risk assessment of the onset), judging the presence or absence of thrombosis or organ damage, Prediction, monitoring, and determination of treatment methods for the prognosis of thrombosis or organ damage. Background technique [0002] DIC is s...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/37G01N33/573
CPCC12Q1/56G01N2800/226G01N33/573G01N2333/96463Y10S436/811G01N2333/968Y10T436/25375G01N33/86G01N2333/974G01N2333/966
Inventor 小野智子
Owner MITSUBISHI KAGAKA IATRON INC
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