Acellular pertussis vaccine detoxified with hydrogen peroxide and preparation method thereof

A technology of hydrogen peroxide and whooping cough, applied in chemical instruments and methods, antibacterial drugs, bacterial antigen components, etc., can solve the problems of difficult sterilization and filtration, complicated reaction, lack of anti-aggregation agents, etc., and achieve good immunogenicity, The effect of high antigen recovery rate and broad-spectrum antigenicity

Inactive Publication Date: 2015-12-02
HUALAN BIOLOGICAL ENG INC +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the detoxification process of this method, EDTA will also be chelated with metal salts, and the reaction is complicated and difficult to control; at the same time, due to the lack of anti-aggregation agents, proteins are not easy to sterilize and filter due to hydrophobic interaction during the detoxification process, and the yield is low, so it is not suitable for large-scale production. Large-scale industrial production

Method used

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  • Acellular pertussis vaccine detoxified with hydrogen peroxide and preparation method thereof
  • Acellular pertussis vaccine detoxified with hydrogen peroxide and preparation method thereof
  • Acellular pertussis vaccine detoxified with hydrogen peroxide and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0069] Example 1: Acquisition of pertussis antigens

[0070] After opening the pertussis phase I strain (No. 58003) purchased from the China Institute for the Control of Pharmaceutical and Biological Products, it was inoculated on the modified bag-ginger medium, and cultivated at 35 to 37 ° C for 70 hours to 72 hours. The second generation and the third generation were inoculated on activated carbon semi-synthetic solid medium, cultured at 35-37°C for 23-24 hours, and then inoculated on FMCL (Frohlich modified cyclodextrin liquid) medium or S-S (Stainer-Scholte) medium, at 35-37 Cultivated for 24 hours to 48 hours for production.

[0071] The static culture method or the fermenter culture method can be used, the temperature is 35.5-36.0 ℃, and the culture process should be sampled for pure bacteria inspection.

[0072] Cultures were harvested after the logarithmic growth phase or before the stationary phase.

[0073] Continuous flow centrifugation was used to separate the cu...

Embodiment 2

[0101] Example 2: Detoxification of PT antigens

[0102] a) The PT antigen purified in Example 1 was replaced by phosphate buffer by means of dialysis or ultrafiltration, with a concentration of 100 mmol / L and a pH of 8.0.

[0103] b) Dilute the antigen after replacing the buffer system in step a to 250 μg / ml.

[0104] c) Add glycerol and urea to the antigen diluted in step b so that the final concentration of glycerol reaches 30% (V / V) and the final concentration of urea reaches 2 mol / L.

[0105] d) After step c, add hydrogen peroxide with a final concentration of 1% (W / V) to adjust the pH to 8.0.

[0106] e) The detoxification temperature is 37°C, and the detoxification time is 4 hours.

[0107] f) Remove hydrogen peroxide by ultrafiltration to terminate detoxification.

[0108] g) After terminating the detoxification in step f, filter with a 0.22 μm filter membrane to obtain the PT stock solution after detoxification.

[0109] The test results of the PT stock solution a...

Embodiment 3

[0112] Example 3: Detoxification of FHA antigens

[0113] a) Replace the purified FHA antigen with phosphate buffer by means of dialysis or ultrafiltration, with a concentration of 100 mmol / L and a pH of 8.0.

[0114] b) Dilute the antigen after replacing the buffer system in step a to 250 μg / ml.

[0115] c) Add glycerol and urea to the antigen diluted in step b so that the final concentration of glycerol reaches 30% (V / V) and the final concentration of urea reaches 2 mol / L.

[0116] d) After step c, add hydrogen peroxide with a final concentration of 0.5% (W / V) to adjust the pH to 8.0.

[0117] e) The detoxification temperature is 37°C, and the detoxification time is 2 hours.

[0118] f) Remove hydrogen peroxide by ultrafiltration to terminate detoxification.

[0119] g) After terminating the detoxification in step f, filter with a 0.22 μm filter membrane to obtain the detoxified FHA stock solution.

[0120] The test results of FHA stock solution after detoxification are ...

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Abstract

The invention relates to an acellular pertussis vaccine, which contains a PT antigen and an FHA antigen in the supernatant part of a bordetella pertussis liquid culture and a PRN antigen in the thallus part of the bordetella pertussis liquid culture. Specifically, the PT antigen and the FHA antigen adopt hydrogen peroxide as the detoxifier, take glycerin as the anti-aggregation agent, and employ urea as the protective agent to conduct detoxification. The detoxified pertussis stock solution has good immunogenicity, is free of residual substances harmful to the human body, and has good immunogenicity and a broader spectrum of antigenicity. The invention also relates to a preparation method of the acellular pertussis vaccine, and a preparation method of a combined vaccine containing the acellular pertussis vaccine.

Description

technical field [0001] The invention relates to a method for detoxifying pertussis vaccine, in particular to a method for detoxifying acellular pertussis vaccine antigen by using hydrogen peroxide. Background technique [0002] Pertussis (pertussis) is a highly contagious severe acute respiratory infectious disease caused by Bordetella pertussis. One of the major infectious diseases of human health. [0003] The causative bacterium of pertussis is B. pertussis in the genus Bordetella, often called B. pertussis. B. pertussis can produce many virulence factors, which can change in phenotype due to changes in environmental conditions, and the expression levels of virulence factors are also different. These factors include pertussis exotoxin (PT); filamentous hemagglutinin (FHA); pertussis Bacillus adhesin (pertactinPRN); thermostable endotoxin (ET); thermolabile toxin (HLT); tracheal cytotoxin (TCT); adenosyl cyclase toxin (ACT) and other biologically active substances. When...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/116A61K39/10A61K39/08A61K39/05A61P31/04C07K14/235
Inventor 苏琴安文琪马小伟张勇朝潘若文范蓓
Owner HUALAN BIOLOGICAL ENG INC
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