Antibody type biological magnetic microsphere as well as preparation method and application thereof

A technology of magnetic microspheres and body shape, applied in the field of biochemistry, can solve the problems of reduced binding performance of purification media, large labor and time, and reduced purification effect, so as to increase the capture rate and binding amount, high efficiency and high yield, Efficient elution effect

Pending Publication Date: 2021-10-29
KANGMA SHANGHAI BIOTECH LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

After the purification column is used many times, the binding performance of the purification medium will be reduced, and the purification effect will be reduced
Therefore, in order to ensure high purification efficiency and quality, operators need to replace all the fillers in the affinity chromatography column in time. This process not only consumes a lot of consumables, but also consumes a lot of labor and time, resulting in high purification costs.

Method used

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  • Antibody type biological magnetic microsphere as well as preparation method and application thereof
  • Antibody type biological magnetic microsphere as well as preparation method and application thereof
  • Antibody type biological magnetic microsphere as well as preparation method and application thereof

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preparation example Construction

[0308] 2. The present invention also discloses a method for preparing biomagnetic microspheres described in the first aspect, comprising the following steps: (i) providing magnetic microspheres combined with biotin or biotin analogs (also known as biotin magnetic microspheres) (biotin ball or biotin magnetic bead); (ii) linking the raw material providing the antibody-type label with the biotin analog at the end of the polymer branch of the biomagnetic microsphere to obtain the biomagnetic microsphere combined with the antibody-type label K (antibody magnetic beads).

[0309] In some preferred modes, the raw material for providing the antibody-type tag is the antibody-type tag.

[0310] In some preferred modes, the raw material for providing the antibody-type tag is a covalently linked complex of avidin or an avidin analogue and the antibody-type tag. In step (ii), the covalently linked complex of avidin or avidin analog and the antibody-type tag is bound to the end of the pol...

Embodiment approach

[0357] Add biotin-modified biomagnetic microspheres D to a fusion protein solution of an avidin-antibody-type label-linked complex (for example, antiEGFP-mScarlet-Tamvavidin2, an antibody-type fusion protein, and a fusion protein of a nanobody) , mix and incubate. Through the specific binding of avidin (such as Streptavidin or Tamvavidin2) to biotin, the antibody-type label is fixed to the end group of the polymer branched chain on the outer surface of the biomagnetic microsphere D, and an avidin-antibody is obtained. Type-labeled biomagnetic microspheres K. Wherein, the fusion protein of avidin-antibody tag can be obtained by in vitro cell-free protein synthesis through IVTT reaction. At this time, the supernatant obtained after the reaction between the biomagnetic microsphere D and IVTT is mixed, and the antibody-type label is realized through the specific binding between the biotin on the outer surface of the biomagnetic microsphere D and the avidin fusion protein in the s...

Embodiment 1

[0384] The preparation of embodiment 1 biomagnetic microsphere D (in conjunction with biotin)

[0385] Preparation of silica-coated magnetic microspheres (also known as magnetic microsphere bodies, magnetic beads, glass beads)

[0386] 20g Fe 3 o 4 The microspheres were placed in a mixed solvent of 310mL ethanol and 125mL water, 45mL 28% (wt) ammonia water was added, 22.5mL tetraethyl orthosilicate was added dropwise, and the reaction was stirred at room temperature for 24h, and washed with ethanol and water after the reaction. Ferric oxide microspheres with different particle sizes (about 1 μm, 10 μm, 100 μm) are used as raw materials to control the particle size of the obtained glass beads. The ferric oxide microspheres with different particle sizes can be prepared by conventional technical means.

[0387] The prepared magnetic microspheres are used as the basic raw material for modifying the purification medium or linking elements-purification medium, so they are also ca...

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Abstract

The invention discloses an antibody type biological magnetic microsphere as well as a preparation method and application thereof. On one hand, the invention provides the biological magnetic microsphere, at least one polymer with a linear main chain and branched chains is fixed onto the outer surface of a magnetic microsphere body, and tail ends of the branched chains of the polymer of the biological magnetic microsphere are connected with antibody type labels. The invention also provides a preparation method and application of the biological magnetic microsphere provided by the invention. The biological magnetic microsphere provided by the invention is convenient in operation and use and can be rapidly dispersed and rapidly settled in a solution, and large-scale experimental facilities such as a high-speed centrifugal machine are not required to be used; the biological magnetic microsphere is an antibody magnetic bead, and the antibody type labels can also be connected to the tail ends of the branched chains of the polymer through interaction of an affinity compound; and the use is wide, and the antibody type labels as a purification medium have flexible selectivity, and are universally and massively applied to separation and purification of a target object.

Description

technical field [0001] The invention belongs to the technical field of biochemistry, and in particular relates to an antibody-type biomagnetic microsphere and a preparation method and application thereof. Background technique [0002] The separation and purification of protein substances is an important downstream link in the production process of biopharmaceuticals. The effect and efficiency of separation and purification directly affect the quality and production cost of protein drugs. For protein purification, materials such as agarose gel are commonly used as purification columns or purification microsphere carriers at this stage. The three-dimensional porous structure of the gel-like material is conducive to increasing the specific surface area of ​​the material, thereby increasing the sites that can bind to the purification medium and increasing the specific binding amount to the target protein. Although the three-dimensional porous structure of the carrier material c...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P21/00C07K1/14
CPCC12P21/00C07K1/14
Inventor 郭敏吴亮徐丽琼曹平生唐磊于雪
Owner KANGMA SHANGHAI BIOTECH LTD
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