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Method for rapidly purifying and isolating anti-actin F(ab')2 and Fc fragments

A technology for purification and separation of proteins, applied in chemical instruments and methods, peptide preparation methods, immunoglobulins, etc., can solve the problems of increased experimental costs, more reagent consumables, and the impact of experimental analysis, so as to reduce experimental costs and instrument maintenance The effect of cost reduction, reduced experiment use time, and simple operation process

Inactive Publication Date: 2017-05-31
GUANGDONG ANNPO BIOTECHNOLOGY INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] There are following shortcoming in the method in the prior art: (1) time-consuming long
(2) High experimental cost
The laboratory needs to be equipped with purification systems, ultra-high-speed centrifuges, ultrafiltration devices or solution replacement devices and other experimental instruments. Experiments use mobile phases, ultrafiltration tubes, protein A affinity chromatography fillers and other reagent consumables, which greatly increase the cost of experiments.
(3) Large consumption of samples and reagents
(4) High requirements for experimenters
(5) Recovery rate and final sample concentration are difficult to control
After purification and separation and subsequent ultrafiltration and concentration, due to the large number of sample transfers, it is very easy to reduce the recovery rate of the sample during processing, which directly leads to the difficulty of confirming the final sample concentration, which may affect the subsequent experimental analysis.

Method used

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  • Method for rapidly purifying and isolating anti-actin F(ab')2 and Fc fragments
  • Method for rapidly purifying and isolating anti-actin F(ab')2 and Fc fragments
  • Method for rapidly purifying and isolating anti-actin F(ab')2 and Fc fragments

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Embodiment

[0042] This protocol allows samples to be obtained within 3 hours and subsequent assays can be performed immediately, and antibody fragments are prepared according to the protocol. 12% separation gel was used to analyze and verify the samples after enzymatic hydrolysis and the samples separated by magnetic beads three times. figure 2 SDS-PAGE results of antibody fragments. In the figure, Lane1 is molecular weight marker (Thermo), Lane2 is endonuclease, Lane3 is intact IgG, Lane4 is digested IgG, Lane5, 7, and 9 are F(ab') 2 Fragments, Lane6, 8, 10 are Fc fragments. Comparing Lane5 and Lane 6, it can be seen that the separation effect of this scheme is really reliable, F(ab') 2 Fragments can be completely separated from Fc fragments. At the same time, F(ab') can be seen by repeated use of magnetic beads 2 The separation effect is still ideal (Lane5, 7, 9), while the Fc fragments are all captured by the magnetic beads (Lane6, 8, 10).

[0043] It can be seen that this metho...

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Abstract

The invention provides a method for rapidly purifying and isolating anti-actin F(ab')2 and Fc fragments, which comprises a proteolysis step and a protein A magnetic bead purification step. The method provided by the invention is simple and convenient to operate, high in purification efficiency, accurate and reliable.

Description

technical field [0001] The invention belongs to the field of biopharmaceuticals, in particular to a purified isolated protein antibody F(ab') 2 and Fc fragment methods. Background technique [0002] In the field of biopharmaceuticals, monoclonal antibody drugs include homologous monoclonal antibodies and recombinant monoclonal antibodies. In recombinant monoclonal antibodies, the F(ab') 2 The Fc fragment and the Fc fragment are derived from two different species, and the two fragments are recombined by genetic engineering and then cultured to produce antibody protein. In the protein antibody structure analysis, because F(ab') 2 The structure and function analysis of each fragment has important guiding significance for the judgment and analysis of the structure and function of the whole protein. For example, due to protein post-translational modification can cause F(ab') 2 The difference between the type of oligosaccharide and the type of oligosaccharide on the Fc fragment...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P21/06C07K16/00C07K1/14
CPCC07K1/14C07K16/00C12P21/06
Inventor 朱晓琦
Owner GUANGDONG ANNPO BIOTECHNOLOGY INC
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