Immobilized metal ion affinity chromatography filler, chromatographic column and preparation method thereof

Abstract

The invention discloses an immobilized metal ion affinity chromatography filler, a chromatographic column and a preparation method thereof. The method comprises the following steps: mixing and stirring potash water glass, gamma-glycidyl ether oxypropyl trimethoxy silane and water-dissolved adenosine disodium triphosphate, then adding water-dissolved formamide, stirring to obtain a reaction solution, reacting and curing to obtain a solid material, and then washing to obtain the immobilized metal ion affinity chromatographic column filler. The ATP modified immobilized metal ion affinity capillary monolithic column prepared through a one-step reaction method is simple and rapid in preparation step, high in repeatability and yield and low in preparation cost. The product can be obtained by uniformly mixing the raw materials and then carrying out one-time baking reaction, and is good in physical and chemical stability, and has relatively good reproducibility after being used for multiple times.

Description

technical field [0001] The invention belongs to the field of chromatography, and in particular relates to an immobilized metal ion affinity chromatography filler, a chromatography column and a preparation method thereof. Background technique [0002] Immobilized metal ion affinity chromatography (IMAC) is used for affinity purification of proteins, enzymes, polypeptides and amino acids that have the ability to bind to metal ions. The principle is to enrich and purify specific proteins / peptides by utilizing the electrostatic interaction between metal ions on solid-phase carriers and some amino acids or chemical modification groups exposed by proteins / peptides. After 30 years of development , has gradually become one of the most effective techniques for separating and purifying biomolecules such as proteins. [0003] The core of immobilized metal ion affinity chromatography (IMAC) lies in the selection and preparation process of solid phase matrix, ligands and metal ions. The...

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Problems solved by technology

[0004] At present, the solid phase matrix of commercial IMAC products is usually resin microspheres or magnetic particles, so that the process of loading, washing and elution can be carried out by means of centrifugation or magnetic adsorption. Iminodiacetic acid (IDA), aminotriacetic acid (NTA ) and ethylenediamine (TED) are representative chelating ligands for immobilizing metal ions, and their carboxyl and amine groups can bind Al 3+ , Fe 3 + , Ga 3+ 、Ni 2+ and other metal ions are chelated on the surface of the solid phase matrix, but the chelation performance of these combinations is poor and the selectivity is not high

At present, whether it is commercialized or reported in the literature, IMAC materials are usually in the form of microspheres or magnetic materials. The enrichment of target proteins and polypeptides is completed in centrifuge tubes. The processes of loading, washing, and elution need to be done manually. It is time-consuming and labor-intensive. In addition, the current commercialized IMAC materials are monopolized by foreign companies, and the price is relatively expensive (for example: Thermo Fisherman's product number A32992, 4629 yuan 30 times)

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