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Purification method for large-scale production of embryogenic antigens

A purification method and antigen technology, applied in the field of vaccine antigen preparation, to achieve the effect of improving the purification effect

Inactive Publication Date: 2020-06-19
YEBIO BIOENG OF QINGDAO
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Aiming at the fact that the existing technology cannot completely solve the problem of denatured impurities remaining in the inactivated antigen solution, the present invention provides a large-scale production method for purifying embryotoxin antigens, which can purify the remaining denatured impurities in the inactivated antigen solution. Completely remove the protein to further improve the safety of the vaccine

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  • Purification method for large-scale production of embryogenic antigens
  • Purification method for large-scale production of embryogenic antigens
  • Purification method for large-scale production of embryogenic antigens

Examples

Experimental program
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Effect test

Embodiment 1

[0027] Example 1: Establishment method

[0028] The method established by the present invention comprises the following steps:

[0029] 1.1 Variable temperature treatment:

[0030] Select the inactivated antigen of the avian influenza virus H5N1 subtype Re-8 strain, the HA is 10Log2, and the turbidity OD600 value is 0.604, and it is rapidly lowered from the inactivated state of 36-37°C to 2-15°C.

[0031] Set up constant temperature, direct stainless steel mesh filter control group.

[0032] 1.2 Stand still:

[0033] Place the inactivated antigen at 2-15°C for 7-14 days. The purpose is to completely separate out the remaining miscellaneous proteins and keep them stable.

[0034] The selection of the standing date not only takes into account the removal efficiency of foreign proteins, but also avoids the loss of the target antigenic protein ( figure 1 ).

[0035] 1.3 High-speed centrifugation:

[0036] The inactivated antigen solution treated with variable temperature is...

Embodiment 2

[0039] Embodiment 2: large-scale production

[0040] According to the method established in Example 1, the avian influenza virus H5N1 subtype Re-8 strain antigen was selected as the research object in the avian influenza inactivated vaccine workshop, and 9 batches were tried continuously, and each batch was sampled before and after the antigen was purified, and the turbidity was monitored. (OD600) and HA (see image 3 , Figure 4 ). From the monitoring data before and after centrifugation at variable temperature of 9 batches of inactivated antigens, the OD600 value dropped from 0.5 to 0.7 before centrifugation to 0.3 to 0.4 after centrifugation. There was no significant change in HA titer; indicating that the method of the present invention can be used for large-scale production applications.

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Abstract

The invention provides a purification method for large-scale production of embryogenic antigens. According to the purification method, residual denatured impure proteins in inactivated antigen liquidcan be thoroughly cleared, so that the safety of a vaccine is further improved. The method comprises the following steps: carrying out cooling during inactivation of embryogenic inactivated antigens by formaldehyde; and carrying out standing, carrying out high speed centrifugation on the inactivated antigens in a centrifugal force of 13000g-16000g, so as to obtain supernate, namely the purified embryogenic antigens. According to the method, the residual impure proteins are denatured and separated through temperature varying and then are removed by virtue of a centrifugation method, and meanwhile, a complete structure of a target antigen is maintained.

Description

technical field [0001] The invention belongs to the technical field of preparation of vaccine antigens, and in particular relates to a purification method for large-scale production of embryotoxin antigens, that is, a purification method for formaldehyde-inactivated embryotoxin antigens under large-scale production conditions. [0002] technical background [0003] Under the current large-scale production conditions, although the embryotoxin antigens including avian influenza virus have undergone the existing centrifugation, concentration and purification processes before formaldehyde inactivation to remove some impurities, the antigen solution remains after formaldehyde inactivation. Miscellaneous protein structure changes, resulting in denaturation. [0004] In the field of existing vaccine preparation, in the preparation process of some vaccines, after formaldehyde inactivation, it will directly enter the follow-up oil emulsion inactivated vaccine emulsification link; ther...

Claims

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Application Information

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IPC IPC(8): C07K14/125C07K14/11C07K14/165C07K14/075C07K1/14
CPCC07K14/005C12N2760/18122C12N2760/16122C12N2710/10122C12N2770/20022
Inventor 韩建文夏娜唐振李龙朱艳梅丁欣李晓林丁财辉于国营杜元钊
Owner YEBIO BIOENG OF QINGDAO
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