Human fibrin in-vitro degradation method

A technology for fibrin and degradation products, applied in the field of in vitro degradation of human fibrin, can solve the problems of FDP and D-dimer purity and stability, complex methods, and inability to be used as quality control or standard products, etc.

Active Publication Date: 2020-01-14
BEIJING SICCEEDER TECH CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] However, in the prior art, most of the FDP and D-dimer are prepared separately to prepare the kit, and this method is more complicated.
And a small amount of preparation of FDP and D-dimer at the same time, and used for detection, but the resulting FDP and D-dimer purity and stability have certain problems, can not be used as quality control or standard

Method used

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  • Human fibrin in-vitro degradation method
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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Embodiment 1 preparation process screening

[0040] 1. Raw material preparation and preparation

[0041] 1.1. Tris (Tris) sodium chloride degradation buffer: 50 mM Tris-HCl, 0.9% normal saline, 0.1% sodium benzoate, pH 8.5.

[0042] 1.2, 2× freeze-drying buffer: 4-hydroxyethylpiperazineethanesulfonic acid (Hepes) 4%, trehalose 1g / L, bovine albumin 70g / L, mannitol 40g / L, sodium benzoate 0.2wt%, pH 7.6-7.8;

[0043] 1 × lyophilization buffer: 4-hydroxyethylpiperazineethanesulfonic acid (Hepes) 2%, trehalose 0.5g / L, bovine albumin 35g / L, mannitol 20g / L, sodium benzoate 0.1wt%, pH 7.6-7.8;

[0044] 1.3, calcium chloride (CaCl 2 ) solution: 25mM CaCl 2 solution.

[0045] 1.4. Fibrinolytic enzyme: Fibrinolytic enzyme purchased from Sigma, reconstituted in pure water, frozen in aliquots and stored in a -20°C refrigerator.

[0046] 1.5. Thrombin: dissolved in normal saline to prepare 120U / ml. Add thrombin to a final concentration of 2-3U / mL each time.

[0047] 1.6, pla...

Embodiment 2

[0084] According to the result of embodiment 1, select following steps to degrade:

[0085] 1. Raw material preparation and preparation

[0086] 1.1 Tris (Tris) sodium chloride degradation buffer: 50 mM Tris-HCl, 0.9% normal saline, 0.1% sodium benzoate, pH 8.5.

[0087] 1.2 2× freeze-drying buffer: 4-hydroxyethylpiperazineethanesulfonic acid (Hepes) 4%, trehalose 1g / L, bovine albumin 70g / L, mannitol 40g / L, sodium benzoate 0.2wt%, pH 7.6-7.8;

[0088] 1 × lyophilization buffer: 4-hydroxyethylpiperazineethanesulfonic acid (Hepes) 2%, trehalose 0.5g / L, bovine albumin 35g / L, mannitol 20g / L, sodium benzoate 0.1wt%, pH 7.6-7.8;

[0089] 1.3 Calcium chloride (CaCl 2 ) solution: 25mM CaCl 2 solution.

[0090] 1.4 Plasmin: Plasmin purchased from Sigma, reconstituted in pure water, frozen in aliquots and stored in a -20°C refrigerator.

[0091] 1.5 Thrombin: dissolved in normal saline to make 120U / ml. Add thrombin to a final concentration of 2-3U / mL each time.

[0092] 1.6 Pla...

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Abstract

The invention relates to the technical field of biochemical detection, in particular to an in-vitro degradation method of human fibrin. According to the invention, fibrins are pretreated in a filtering and grinding mode, the prepared fibrin forms uniform and fine particles under the action of mechanical shearing force, and subsequent plasmin degradation is facilitated, by controlling the adding amount of plasmin and rapidly centrifuging after fibrinolysis is finished to obtain a supernate, and a freeze-dried product prepared by adding a certain amount of a freeze-drying formula can be stably stored for more than one year for a long time. In addition, the experiments prove that more stable degradation products can be obtained under the parameters in the scheme, and the method has more obvious advantages compared with other parameters.

Description

technical field [0001] The invention relates to the technical field of biochemical detection, in particular to a method for degrading human fibrin in vitro. Background technique [0002] The fibrinolytic system, referred to as the fibrinolytic system, refers to the transformation of plasminogen into plasmin under the action of plasmin. The fibrinolytic system mainly includes fibrinolytic activators, inhibitors and fibrinolytic proteins. The activation of the fibrinolytic system in the body mainly includes three pathways, namely, the internal activation pathway, the external activation pathway and the exogenous activation pathway. The internal activation is mainly initiated by related factors of the endogenous blood coagulation system, factors XIIa, XIa, high molecular weight kininogen HMWK , the participation of kallikrein is the theoretical basis of secondary fibrinolysis; the external activation is mainly the effect of tissue plasminogen activator (t-PA) and urokinin plas...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P21/06G01N33/96
CPCC12P21/06G01N33/96
Inventor 丁重辉李博华胡晓娟杨娟
Owner BEIJING SICCEEDER TECH CO LTD
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