Preparation method of recombinant protein A mediated immune targeting nano-gold

A recombinant protein, nano-gold technology, applied in the fields of nanotechnology, nanotechnology, nanotechnology for materials and surface science, can solve the problems of low antibody loading rate and biological activity, and inability to form disulfide bonds, and achieve good results. Antibody orientation effect, increase binding capacity, and effect of good application prospects

Active Publication Date: 2017-07-07
天津市泌尿外科研究所 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

In addition, due to the lack of cystine and cysteine ​​in ProteinA\G, disulfide bonds cannot be formed, which ensures that ProteinA\G can specifically bind to the Fc segment of the antibody to achieve better antibody orientation than physical adsorption and chemical bonding. , which is expected to be the key to solving the problem of low loading rate and biological activity of immune-targeted gold nanoparticles

Method used

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  • Preparation method of recombinant protein A mediated immune targeting nano-gold
  • Preparation method of recombinant protein A mediated immune targeting nano-gold
  • Preparation method of recombinant protein A mediated immune targeting nano-gold

Examples

Experimental program
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Effect test

Embodiment 1

[0039] Example 1: Recombinant ProteinA\G-mediated 20nm nano-gold linking with SPACA3 antibody to prepare immuno-targeted nano-gold

[0040] The preparation method of immune targeting nano-gold system, the steps are as follows:

[0041] The antibody Fc segment forms a targeted immune nano-gold system by specifically binding to the recombinant ProteinA\G on the surface of nano-gold. The preparation method of the immune-targeted nano-gold includes the following steps:

[0042] 1. Preparation of 20nm nano-gold: In the process of preparing nano-gold by trisodium citrate reduction method, all glass instruments are cleaned with newly configured aqua regia (HNO3:HCL, 1:3), rinsed with double distilled water, and baked before use dry. Add 50 ml of 0.01% chloroauric acid to an Erlenmeyer flask and place it on a heating magnetic stirrer. The fixed reaction temperature is 95° C., and the stirring speed is 300 r / min. Quickly add 0.75ml of 1% trisodium citrate to the above solution. After 25s, t...

Embodiment 2

[0082] Example 2: Recombinant ProteinA\G-mediated 40nm nano-gold connected to SPAG8 antibody to prepare immuno-targeted nano-gold;

[0083] The preparation method of immune targeting nano-gold system, the steps are as follows:

[0084] The antibody Fc segment forms a targeted immune nano-gold system by specifically binding to the recombinant ProteinA\G on the surface of nano-gold. The preparation method of the immune-targeted nano-gold includes the following steps:

[0085] 1. Preparation of 40nm nano-gold: In the process of preparing nano-gold by trisodium citrate reduction method, all glass instruments are cleaned with newly configured aqua regia (HNO3:HCL, 1:3), rinsed with double distilled water, and baked before use dry. Add 100 ml of 0.01% chloroauric acid to an Erlenmeyer flask and place it on a heating magnetic stirrer. The fixed reaction temperature is 120° C., and the stirring speed is 400 r / min. Quickly add 1ml of 1% trisodium citrate to the above solution. After 25s, th...

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Abstract

The invention provides a preparation method of recombinant protein A mediated immune targeting nano-gold. A recombinant Protein A / G is used as an affinity ligand to enable an antibody fix on the surface of the nano-gold to form an immune nano-gold compound. The compound is composed of three parts including the antibody, the recombinant protein A and the nano-gold; the recombinant protein A contains C-terminal cysteine after being modified and can be coupled on the nano-gold through a single site, so that steric hindrance is reduced and a binding capacity between the recombinant protein A and the antibody is also increased; an Fc segment of the antibody is specifically combined through a non-covalent bond and an antibody orientation effect which is better than physical adsorption and chemical bonding is obtained. The method can be used for preparing complicated multi-component and multifunctional compound immune targeting nano-gold to meet different requirements. The immune targeting nano-gold prepared by the invention not only has reaction specificity and stability, but also has relatively high anti-interference capability and environment tolerance and has a very good application prospect in the fields including targeting tracing, biological pharmacy, accurate treatment and the like.

Description

Technical field [0001] The present invention relates to the coupling technology of nano-gold and antibody, in particular to the preparation of immuno-targeted nano-gold by using antibody Fc section and the recombinant protein A (ProteinA\G) on the surface of self-assembled nano-gold to specifically combine. Background technique [0002] In recent years, with the development of nanotechnology, the application of nanomaterials has gradually become the focus of research. As an important member of metal nanomaterials, nano-gold, in addition to its own unique physical and chemical properties, also has properties such as small size effect, surface effect and macroscopic quantum tunneling effect. Therefore, nano-gold is used in substance detection, biosensor and immune targeting, etc. Shows great potential in the field. Especially in the field of immunotargeting, as a stable, labelable, and highly biosafe carrier, gold nanoparticles need to be combined with targeted substances to obtai...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/543B82Y30/00
CPCB82Y30/00G01N33/54346
Inventor 王江牛远杰王荷芳李刚沈丽琳蒋宁尚芝群王准陈家童
Owner 天津市泌尿外科研究所
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