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Protein A immuo adsorption material and preparing method

An immunoadsorbent material and protein technology, which is applied in the field of protein A immunoadsorbent material for blood purification and its preparation, can solve the problems of large batch-to-batch variation, many reaction steps, unstable performance and the like of the adsorbent material product, so as to improve the binding efficiency. , The preparation process is simplified, and the regeneration performance is good.

Active Publication Date: 2007-11-14
GUANGZHOU KONCEN BIOSCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although these two methods eliminate the highly toxic substance cyanogen bromide, and the materials prepared by them are safer to use, but there are many reaction steps in the preparation process, and five steps of chemical reactions are required to synthesize the adsorption material. The method is complicated, so the obtained adsorption There are large differences between batches of materials and products, and the performance is unstable

Method used

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  • Protein A immuo adsorption material and preparing method
  • Protein A immuo adsorption material and preparing method
  • Protein A immuo adsorption material and preparing method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Synthesis of Activated Sepharose Containing Epoxy Groups (Sepharose 6FF)

[0038] 1. Reaction of agarose gel with ethylene glycol bisglycidyl ether

[0039] Add 1.5 liters of agarose gel (Sepharose 6FF) in a 5L reactor, 1000ml of 0.8mol / L NaOH aqueous solution, mix well, add 1000ml of ethylene glycol bisglycidyl ether, place in a constant temperature shaker, at 20 °C for 2 hours. After the reaction was completed, the gel was filtered and rinsed with a large amount of distilled water until neutral. Store the activated medium at 4°C for later use. The number of epoxy groups in the gel activated by this method was detected by the sodium thiosulfate method, and it was measured that there were at least 45 μmol of epoxy active groups per milliliter.

[0040] 2. Reaction of Sepharose with Butanediol Bisglycidyl Ether

[0041] Add 1 liter of agarose gel (Sepharose 6FF) and 1000ml of 1mol / L NaOH aqueous solution into a 5L reactor, mix well, add 1000ml of butanediol diglycidy...

Embodiment 2

[0047] Synthesis of Protein A Immunosorbent Material

[0048] 1. Activated agarose gel (Sepharose 6FF) reacted with protein A to synthesize immunoadsorbent material

[0049] In the 5L reactor, add 1 liter of epoxy-active agarose gel (Sepharose 6FF) synthesized in Example 1 (1~3), 1.5L of 0.1mol / L boric acid buffer solution, and the pH value is controlled in the range of 8.0~10.0 Inside, add 6.0g of protein A, react at a constant temperature of 30°C for 16 hours, stop the reaction, recover unreacted protein A, cap the unreacted epoxy active group with 0.2mol / L glycine ethyl ester hydrochloride, and React at room temperature for 5 hours. Rinse the unreacted components with a large amount of distilled water, store them in 0.1mol / L phosphate buffer solution with pH=7.4, and add preservatives for storage. The binding amount of protein A detected by ultraviolet method was 5-5.5 mg / ml.

[0050] 2. Activated agarose gel (Sepharose 6FF) reacts with protein A to synthesize an immunos...

Embodiment 3

[0052] Animal Test of Protein A Immunosorbent Material

[0053] Put 20ml of the protein A immunosorbent material synthesized in Example 2 (1), in which the binding amount of protein A is 100mg, put it into a beaker and soak it with normal saline for 1 hour, then put it in a column of Φ20×50mm, and wash it with normal saline Rinse the column well. Taking the animal dog as the test object, after the dog is anesthetized, the blood is pumped out from the femoral artery of the dog at a speed of 30ml / min, the blood cells and plasma are separated through a plasma separator, and the plasma is pumped into protein A at a speed of 10ml / min The adsorption column is absorbed and then mixed with blood cells and pumped into the dog's femoral vein. After 5 minutes of adsorption, the plasma in the column was washed out with physiological saline until the absorbance at 280 nm was zero. The immunoglobulin and its immune complexes adsorbed in the column were eluted with 0.2 mol / L, pH=2.3 glycin...

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Abstract

The present invention relates to a protein A immunoadsorption material for purifying blood and its preparation method. Said protein A immunoadsorption material is a high-molecular material which is obtained by using agarose gel as carrier matrix, making said agarose gel be reacted with diglycidyl ether coupling reagent to obtain active carrier, then making said active carrier and protein A implement coupling reaction. Said invention can be used for making clinical immunoadsorption therapy.

Description

technical field [0001] The invention belongs to medical biological materials, in particular to a protein A immunoadsorbing material used for blood purification and a preparation method thereof. Background technique [0002] The occurrence and development of many diseases are the result of the accumulation of pathogenic factors in the body. To be able to specifically and effectively remove these pathogenic factors from the body through the method of purifying the blood without causing damage to the body has been a problem that has been continuously explored in clinical medicine for the past ten or twenty years. Immunoadsorption therapy is an important method of blood purification. Its principle is to firmly bind a ligand to a fixed carrier to form an immunoadsorption column, and specifically remove pathogenic substances in the blood of patients through extracorporeal circulation. Factors, purify the blood, so as to achieve the purpose of treating diseases. At present, the c...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61L2/16A61L101/46
Inventor 陈校园汪志刚李光吉许春生张旭锋
Owner GUANGZHOU KONCEN BIOSCI
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