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An affinity monolithic column based on a graphene-nano-gold composite interface with ultra-high loading of nucleic acid aptamers and its preparation method

A nucleic acid aptamer and nano-gold technology, which is applied in chemical instruments and methods, separation methods, solid adsorbent liquid separation, etc., can solve the problem of limited binding sites and difficulties in greatly improving the coverage density of nucleic acid aptamers, etc. question

Active Publication Date: 2021-07-02
XIAMEN HUAXIA UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] 1. Multiple derivatization methods, mainly including non-chemical bonding method and chemical bonding method, through the affinity or chemical coordination between nucleic acid aptamers and the overall surface, repeated post-column derivatization of nucleic acid aptamers to further improve nucleic acid adaptation The coverage density of the body on the monolithic column, in which the non-chemical bonding method is mainly biotin and streptavidin method, and the chemical bonding method mainly includes the thio-ene click and glutaraldehyde method, but due to the limited binding sites, this method It is difficult to greatly improve the coverage density of nucleic acid aptamers

Method used

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  • An affinity monolithic column based on a graphene-nano-gold composite interface with ultra-high loading of nucleic acid aptamers and its preparation method
  • An affinity monolithic column based on a graphene-nano-gold composite interface with ultra-high loading of nucleic acid aptamers and its preparation method
  • An affinity monolithic column based on a graphene-nano-gold composite interface with ultra-high loading of nucleic acid aptamers and its preparation method

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Embodiment 1

[0037] An affinity monolithic column based on a graphene-nano gold composite interface with ultra-high loading of nucleic acid aptamers and a preparation method thereof, the specific steps are:

[0038] (1) Capillary pretreatment:

[0039] Silanol is the basis for pre-polymerization on the inner wall of the capillary, and the more the number of exposed silanol on the tube wall, the more favorable the pre-polymerization. However, the number of silanol groups in ordinary fused silica capillaries is very small, which is not conducive to prepolymerization. Therefore, pretreatment of the capillary is required,

[0040] The preprocessing process is as follows:

[0041] The empty capillary column was rinsed with 1.0 mol / L HCl solution for 30 min in sequence, passed through the secondary water to neutral, rinsed with 1.0 mol / L NaOH solution for 30 min, and then heated at 100°C for 3 h, and then sequentially washed with Pass the secondary water to neutral, rinse with 0.1 M hydrochlo...

Embodiment 2

[0054] Formula B was used to prepare gold nanoparticles modified but not modified anti-ochratoxin A nucleic acid aptamer as a control column and an affinity monolithic column modified anti-ochratoxin A nucleic acid aptamer, and equilibrated, enriched and washed respectively And elution, its specific steps are as follows:

[0055] (1) Equilibration: equilibrate the control column and the modified anti-ochratoxin A nucleic acid aptamer affinity column with binding buffer. Binding buffer: 10 mM Tris-HCl (pH 8.0), 120 mM NaCl, 5 mM KCl, 20 mM CaCl 2 500 psi back pressure valve, flow rate 0.05 mL / min, equilibrate for 0.5 h;

[0056] (2) Enrichment: Inject 20 μL of 5 ng / mL ochratoxin A (OTA) solution, respectively, and enrich on the control column and modified OTA nucleic acid aptamer affinity column for 0.5 h. 500 psi back pressure valve, flow rate 0.05 mL / min, collect the enriched solution, to be tested.

[0057] (3) Cleaning: Wash the monolithic column with binding buffer, af...

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Abstract

The invention belongs to the field of preparation of monolithic column polymer materials, and in particular relates to an affinity monolithic column based on graphene-nano-gold composite interface with ultra-high loading of nucleic acid aptamers and a preparation method thereof. Using surface-modified graphene oxide as a catalyst and modification material, a graphene-functionalized hybrid silica gel polymerization monolithic column was prepared by a "one-pot" method, and the nano-gold post was modified to the graphene material of the modified silica gel porous monolithic column On the surface, based on the bridging effect of gold nanoparticles, the ultra-high density loading of nucleic acid aptamers on the surface of the modified silica porous monolithic column is realized. The modified silica gel porous monolithic column matrix has a high specific surface area, and the modified graphene oxide on its surface can stably and efficiently load nano-gold particles, while avoiding the agglomeration of graphene and nano-gold particles in the preparation process, which can make The aptamer can be modified on the monolithic column at a high density, realizing that the silica gel hybrid affinity monolithic column can be used for efficient and specific recognition and separation of ochratoxin A.

Description

technical field [0001] The invention belongs to the field of preparation of monolithic column polymer materials, and in particular relates to an affinity monolithic column based on a graphene-nano gold composite interface with ultra-high loading of nucleic acid aptamers and a preparation method thereof. Background technique [0002] Nucleic acid aptamer is a short-chain DNA or RNA sequence screened from a large-capacity random oligonucleotide library through in vitro screening technology (SELEX), which can bind to the corresponding target ligand with high affinity and high specificity , has the advantages of a wide range of applicable targets, easy synthesis, easy modification, stable chemical properties and high specificity. The development of nucleic acid aptamers provides a new direction for the research of fast and efficient recognition technology. The research work of using nucleic acid aptamers as affinity ligands for enrichment, separation and detection has received h...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): B01D15/08B01D15/38
CPCB01D15/08B01D15/3819
Inventor 池金鑫黄桂华於霞
Owner XIAMEN HUAXIA UNIV