Protein A adsorption medium

An adsorption medium and protein technology, applied in other chemical processes, chemical instruments and methods, etc., can solve the problem that protein A immunoadsorption column products cannot be finally sterilized

Active Publication Date: 2012-10-03
GUANGZHOU KONCEN BIOSCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to provide a protein A adsorption medium that can withstand autoclaving to solve the problem that protein A immunoadsorption column products cannot be subjected to final product sterilization

Method used

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  • Protein A adsorption medium
  • Protein A adsorption medium
  • Protein A adsorption medium

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Experimental program
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preparation example Construction

[0014] 1. Preparation of recombinant protein A

[0015] Obtain the gene coding sequence of recombinant protein A by the following methods:

[0016] 1. Directly synthesize the full sequence (SEQ ID NO.2);

[0017] 2. Synthesize the coding sequence (SEQ ID NO.3) of the repeating unit constituting the recombinant protein A. There is an AccI restriction site at both ends of the sequence. Since the recognition sequence of AccI is a non-palindromic sequence, it contains repeating units After the DNA fragment of the coding sequence is digested by the enzyme, it can be connected end-to-end to obtain a DNA fragment containing the coding sequence of several repeating units (Nilsson B, et al. A synthetic IgG-binding domain based on staphylococcal protein A. Protein Engineering, vol.1, 1987, p107-113), and then select the gene coding sequence of recombinant protein A with the complete sequence (SEQ ID NO.2).

[0018] While obtaining the recombinant protein A coding sequence, add suitabl...

Embodiment

[0024] Embodiment: the preparation of recombinant protein A

[0025] Directly synthesize the DNA fragment (SEQ ID NO.3) containing the repeating unit coding sequence of recombinant protein A. After digestion with NdeI and XhoI, the above DNA fragment is connected into the vector pUC18, which is well known in the art, and the vector is digested with AccI , the resulting small fragments are connected end to end, and then connected to the carrier fragments to obtain a recombinant plasmid containing several repeating unit coding sequences, and the recombinant protein A coding sequence (SEQ ID NO. 2) for the recombinant plasmid, the recombinant protein A coding sequence was transferred to the prokaryotic expression plasmid pET22b well known in the art by digestion with NdeI and XhoI to obtain the recombinant expression plasmid. The recombinant expression plasmid was transformed into Escherichia coli expression strain BL21(DE3), and the obtained recombinant expression strains were...

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Abstract

The invention relates to a protein A adsorption medium. The protein A adsorption medium is formed of a solid phase carrier material and a ligand fixed on the carrier by chemical coupling, wherein the ligand is recombinant protein A of an amino acid sequence represented by SEQ ID NO.1 in a sequence table. The protein A adsorption medium is tolerable to 20 minutes of thermal pressure sterilization at the temperature of 121.5 DEG C, so that a problem that a protein A immune adsorption column product is unable to perform final sterilization is resolved.

Description

technical field [0001] The invention belongs to the field of genetic engineering technology and antibody adsorption technology, and relates to a protein A adsorption medium. Background technique [0002] Protein A comes from the cell wall of Staphylococcus aureus, and its full name is Staphylococcus aureus protein A, referred to as protein A or SPA. The gene coding sequence of natural protein A consists of 1464 bases, encoding a total of 488 amino acids, with a molecular weight of about 42kd, including six structural domains of E, D, A, B, C and X (from the amino terminal to the carboxyl terminal). As we all know, because protein A can selectively adsorb human or mammalian antibodies (mainly IgG, including some IgA and IgM), it is widely used in many fields such as antibody purification, medical diagnosis and clinical treatment. [0003] In terms of clinical treatment, the protein A adsorption medium with protein A as the ligand is used to treat autoimmune diseases such as ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): B01J20/24B01J20/26
Inventor 陈校园余波光杨正根张海珍
Owner GUANGZHOU KONCEN BIOSCI
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