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Purification and storage method of interleukin 15 and receptor-monoclonal antibody fusion protein thereof

An interleukin and monoclonal antibody technology, which is applied in the preparation methods of peptides, chemical instruments and methods, fusion polypeptides, etc., can solve the problems of unstable recovery rate of antibody fusion proteins and so on.

Pending Publication Date: 2021-12-21
SUZHOU INST FOR BIOMEDICAL RES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] In order to solve the above technical problems, the present invention discloses a method for purifying and storing interleukin-15 and its receptor-monoclonal antibody fusion protein. Solve the problem of instability and low recovery of antibody fusion protein during purification

Method used

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  • Purification and storage method of interleukin 15 and receptor-monoclonal antibody fusion protein thereof
  • Purification and storage method of interleukin 15 and receptor-monoclonal antibody fusion protein thereof
  • Purification and storage method of interleukin 15 and receptor-monoclonal antibody fusion protein thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] 1. Collect interleukin-15 and its receptor-monoclonal antibody fusion protein IL15-IL15Rα-aEGFR supernatant;

[0036] 2. Wash the Protein A affinity filler coupled to 4% agarose with 10 column volumes of phosphate buffered saline (DPBS, pH 7.5), repeating several times to remove the storage solution in Protein A;

[0037] 3. Slowly add the protein supernatant obtained by centrifugation to the chromatography column, and repeat the addition several times to combine the fusion protein with the filler;

[0038] 4. Rinse the Protein A affinity filler with 10 times the column volume of phosphate buffered saline (DPBS, pH 7.5), and repeat it several times to remove weakly bound impurities;

[0039] 5. Add 3 times column volume of eluent (Elution buffer, EB), incubate at room temperature for 10 minutes, elute the fusion protein, repeat several times until the fusion protein is completely eluted; use eluent 1 ( EB1), eluent 2 (EB2) to elute the fusion protein, the specific conf...

Embodiment 2

[0045]Concentrate using a concentrator tube at a centrifuge speed of 4000 rpm. When the volume of the fusion protein is concentrated to less than 100 μL, add the storage solution to be replaced, continue centrifuging at the same speed, and repeat this step 3 times, so that the fusion protein is finally in the storage solution to be replaced. The configuration of the stock solution is shown in Table 2.

[0046] Table 2 Configuration of storage solution

[0047] buffer name formula BEX1 DPBS, pH 6.5 BEX2 0.1M sodium citrate, 0.1M~0.2M sodium chloride, pH 6.0 BEX3 0.1M sodium citrate, 0.1M~0.2M sodium chloride, 250mM sucrose, pH 6.0

[0048] The results after changing the stock solution are shown in Table 3.

[0049] Table 3 purification and recovery results

[0050]

[0051] It can be seen from the above table:

[0052] The yield of the fusion protein eluted with the traditional eluent is 20.72 mg / L. During the elution process, the fusi...

Embodiment 3

[0059] The stock solution was replaced with 0.08M sodium citrate, 0.1M-0.2M sodium chloride and 240mM sucrose, pH 6.0, and the rest of the steps were the same as in Example 2.

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Abstract

The invention discloses a purification and storage method of interleukin 15 and a receptor-monoclonal antibody fusion protein thereof. The interleukin 15 and the receptor-monoclonal antibody fusion protein thereof are eluted by using an eluent, the purified fusion protein is stored in a storage solution, and components of the eluent and the storage solution are specifically disclosed. According to the invention, the eluent and the storage solution for the interleukin 15 and the receptor-monoclonal antibody fusion protein IL15-IL15R alpha-aEGFR thereof are optimized, a category of stable buffer solution components are found, and the recovery rate and the purity of the protein are remarkably improved.

Description

technical field [0001] The invention relates to the field of antibody separation and purification, in particular to a method for purifying and storing interleukin-15 and its receptor-monoclonal antibody fusion protein. Background technique [0002] Interleukin-15 (IL-15) is a cytokine with various biological activities, which can be produced by activated monocyte-macrophages, epidermal cells and fibroblasts and other cells. IL-15 was first discovered because it has a function similar to IL-2, which can stimulate the activation and proliferation of CTLL cells. IL-15 can stimulate T cells to produce various activation antigens, such as CD24, CD122, CD95, CD30 and CD69. However, the role of IL-15 in the development of other immune cells was gradually discovered, such as NK cells, NK-T cells and intestinal intraepithelial lymphocytes. A number of studies have shown that IL-15 plays an important role in reducing inflammation, anti-oxidative stress, regulating lipid metabolism a...

Claims

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Application Information

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IPC IPC(8): C07K19/00C07K1/22C07K1/02
CPCC07K14/5443C07K14/7155C07K16/2863C07K2319/00
Inventor 张雨菡郭超王峰张捷
Owner SUZHOU INST FOR BIOMEDICAL RES
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