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Bionic affinity and purification method of plasminogen activator

A technology of plasminogen and purification method, applied in the biological field, can solve the problems of unstable nature, high cost, limited large-scale application, etc., and achieve the effect of reducing production cost and reducing large-scale purification steps.

Inactive Publication Date: 2007-09-05
上海荣君生物医药科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the D-Phe-D-Phe-Argal material used in this method needs to be synthesized by solid-phase peptide synthesis, and the preparation steps are cumbersome and costly; Stable and cannot withstand the harsh conditions of in-line cleaning
Although the complex purification process can be simplified, the above shortcomings limit large-scale applications

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] 1. Preparation of separation materials

[0018] Take NH 2 -Sepharose (250ml), washed with 5 times the volume of 1M NaCl, then fully washed with deionized water, drained of water, poured into a reaction vessel, added 200ml of deionized water, and placed in an ice-salt bath for precooling. When the temperature dropped to 5°C, start stirring. Triazoxide (45 g) was dissolved in pre-cooled acetone and added to the reaction vessel. with saturated NaHCO 3 Keep the pH of the solution between 6 and 7, react at 5°C for 3 hours, take out, 3×10 times the volume of water / acetone (1:1, 1:3, 0:1, 1:1, 3:1, 1:0) and washed successively to obtain 1-amino-Sepharose-3,5-dichloro-2,4,6-triazoxide (190ml).

[0019] Take 1-amino-Sepharose-3,5-dichloro-2,4,6-triazoxide (200ml), weigh aminobenzamidine (20g) and dissolve it with deionized water (150ml), and dissolve it with 1-amino- Sepharose-3,5-dichloro-2,4,6-triazoxide was mixed evenly, placed at a temperature of 50° C. and stirred for ...

Embodiment 2

[0023] 1. Preparation of separation materials

[0024] Take NH 2 -Sepharose (250ml), washed with 5 times the volume of 1M NaCl, then fully washed with deionized water, drained of water, poured into a reaction vessel, added 200ml of deionized water, and placed in an ice-salt bath for precooling. When the temperature dropped to 5°C, start stirring. Triazoxide (45 g) was dissolved in pre-cooled acetone and added to the reaction vessel. with saturated NaHCO 3 Keep the pH of the solution between 6 and 7, react at 5°C for 3 hours, take out, 3×10 times the volume of water / acetone (1:1, 1:3, 0:1, 1:1, 3:1, 1:0) and washed successively to obtain 1-amino-Sepharose-3,5-dichloro-2,4,6-triazoxide (190ml).

[0025] Take 1-amino-Sepharose-3,5-dichloro-2,4,6-triazoxide (200ml), weigh aminobenzamidine (20g) and dissolve it with deionized water (150ml), and dissolve it with 1-amino- Sepharose-3,5-dichloro-2,4,6-triazoxide was mixed evenly, placed at a temperature of 50° C. and stirred for ...

Embodiment 3

[0029] 1. Preparation of separation materials

[0030] Take NH 2 -Sepharose (250ml), washed with 5 times the volume of 1M NaCl, then fully washed with deionized water, drained of water, poured into a reaction vessel, added 200ml of deionized water, and placed in an ice-salt bath for precooling. When the temperature dropped to 5°C, start stirring. Triazoxide (45 g) was dissolved in pre-cooled acetone and added to the reaction vessel. with saturated NaHCO 3 Keep the pH of the solution between 6 and 7, react at 5°C for 3 hours, take out, 3×10 times the volume of water / acetone (1:1, 1:3, 0:1, 1:1, 3:1, 1:0) and washed successively to obtain 1-amino-Sepharose-3,5-dichloro-2,4,6-triazoxide (190ml).

[0031] Take 1-amino-Sepharose-3,5-dichloro-2,4,6-triazoxide (200ml), weigh aminobenzamidine (20g) and dissolve it with deionized water (150ml), and dissolve it with 1-amino- Sepharose-3,5-dichloro-2,4,6-triazoxide was mixed evenly, placed at a temperature of 50° C. and stirred for ...

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PUM

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Abstract

The invention relates to a plasminogen activator bionics and purifying method. It belongs to biotechnology field. It includes the following steps: basal chromatography medium reacts with tri-chlorine tri-nitrogen zin; they react with pair amino benzene formamidine to compound bionics and separating material; the upper material is made into affinity column; the sample of the plasminogen activator is flowed through the affinity column; then the former is absorbed in the latter; cleaning the latter; cleaning it again after changing buffer solution condition; purging the plasminogen activator to gain a purified one. The advantages of the invention are less purifying steps, low cost, high efficiency, and can realize volume produce.

Description

technical field [0001] The invention relates to a method in the field of biotechnology, in particular to a method for biomimetic affinity purification of plasminogen activator. Background technique [0002] Tissue plasminogen activator is a serine protease that has a high affinity with fibrin and exhibits activity after binding to fibrin in the circulatory system. It can activate plasminogen to become plasmin, dissolve thrombus. Tissue plasminogen activator has a natural form, and there is also a form of artificially designed part of the domain deletion, such as the modified form of the deletion of the fibrin ring region, the epidermal growth factor (E) region and / or the Kringle-1 region, and Prolongs half-life in vivo, speeds up diffusion into and dissolves blood clots. Tissue plasminogen activator is a high-efficiency thrombolytic drug, and it is one of the six major gene recombination drugs on the US market. Commonly used purification methods for plasminogen activators...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N9/50
Inventor 李荣秀吴方庞艳萍
Owner 上海荣君生物医药科技有限公司
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