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Meningococcal vaccine and its preparing method

A meningococcal and combined vaccine technology, which is applied in chemical instruments and methods, antibacterial drugs, pharmaceutical formulations, etc., can solve the problems of high equipment investment requirements, no further separation, and numerous and complicated steps, so as to improve immunogenicity , Reduction of reactogenicity, effect of purification process improvement

Inactive Publication Date: 2003-11-26
BEIJING SANROAD BIOLOGICAL PROD CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the high centrifugation speed required and the need for repeated density gradient centrifugation and dissolution, the equipment investment requirements are high, very cumbersome, and the steps are numerous and complicated
The product obtained was a mixture of various group B meningococcal outer membrane proteins without further isolation

Method used

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  • Meningococcal vaccine and its preparing method
  • Meningococcal vaccine and its preparing method
  • Meningococcal vaccine and its preparing method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0049] Preparation of Group B Meningococcal Outer Membrane Protein

[0050] Kaikai Group B meningococcal strains were inoculated on a semi-comprehensive slope, expanded and transferred to the fifth-generation liquid tank for culture, and the bacteria were collected by continuous centrifugation, and 0.3-0.6M LiCl and 0.2-0.5MCH 3 The COONa mixture was used to extract outer membrane proteins. Shake at 35-45°C for 1-3 hours. Centrifuge at 4000-8000 rpm for 1 hour, collect the supernatant, then add ethanol to 60-80% to precipitate the outer membrane protein. After reconstitution with the injection solution, the 400KD filtrate was collected by ultrafiltration, and then concentrated by a 30KD ultrafiltration membrane. Use SDS-PAGE to detect the content of the desired type 1, 2 or 1, 3 outer membrane protein. Chromatography with Sephacryl S 200-400 to collect the required type 1, 2 or 1, 3 outer membrane proteins. Detection of protein content (Lowry method), main components and t...

Embodiment 2

[0052] Group A meningococcal polysaccharide conjugate

[0053] Group A meningococcus capsular polysaccharide diluted to 4mg / ml, add cyanogen bromide to activate the polysaccharide, act at 23±3℃ for 1-1.5 hours, adjust pH and maintain pH9-11, then add adiphydrazide 2.5-5mg / ml mgps, maintain pH8-10 for 10-30 minutes, stir at 2-8°C for 10-20 hours, dialyze to remove small molecular substances, take samples to measure the residual amount of cyanogen bromide and the derivatization rate of adipic hydrazide. Add an equal amount of group B meningococcal outer membrane protein and polysaccharide to mix, add carbodiimide: 15-25mg / ml mixture, act at 5-15°C for 1-1.5 hours, and adjust pH to 5-7. The coupling stock solution was dialyzed to remove small molecular substances. 300KD membrane bag ultrafiltration is concentrated, through Sepharose CL-4B column chromatography, collects the coupling compound of high molecular weight (see Figure 4 ), sterilizing filtration to detect phosphorus ...

Embodiment 3

[0055] Group C meningococcal capsular polysaccharide conjugate

[0056] Group C meningococcal polysaccharide diluted to 4 mg / ml, add cyanogen bromide (CNBr) to activate the polysaccharide, act at 20-30°C for 1-1.5 hours, adjust pH and maintain pH 9-11, then add adipic hydrazide (ADH) 2.5-5 mg / mg polysaccharide, maintain pH 8-10, react for 30 minutes, stir at 2-8°C for 10-20 hours, dialyze to remove small molecular substances. Sampling was carried out to measure residual CNBr content and ADH derivatization rate. Add an equal amount of group B meningococcal outer membrane protein and polysaccharide to mix, add carbodiimide (EDAC): 15-25mg / ml mixture, react at 5-15°C for 1-1.5 hours, adjust pH to 5-7. The coupling stock solution was dialyzed to remove small molecular substances, concentrated by ultrafiltration in a 300KD membrane bag, and subjected to Sepharose CL-4B column chromatography to collect high molecular weight coupling compounds (see Figure 5 ), sterilizing filtrati...

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Abstract

The present invention relates to meningococcal vaccine and its preparation process. Purified colony-A meningococcus polysaccharide and colony-C meningococcus polysaccharide are made to couple chemically with colony-B meningococcus outer membrane protein separately to form monovalent binders, and the monovalent binders are then mixed to compound the vaccine. The vaccine may be used for infant to prevent epidemic cerebrospinal meningitis caused by colony-A and colony-C meningococcus and ichorrhemia.

Description

technical field [0001] The present invention relates to a meningococcal vaccine. In particular, the invention relates to a meningococcal conjugate vaccine. More specifically, the present invention relates to a conjugated vaccine comprising capsular polysaccharides of group A and C meningococci coupled with outer membrane protein of group B meningococcus. The present invention also relates to the preparation method of this vaccine. Background technique [0002] Illness due to Neisseria meningitidis is an important contributor to severe illness and high mortality worldwide. Before the invention of sulfonamides and antibiotics, the fatality rate of meningococcal disease was about 70%. Today, despite good treatment, the case fatality rate of the disease can be as high as 10% even in developed countries, and the case fatality rate of sepsis caused by it can exceed 50%. Neisseria meningitidis causes 500,000 cases and 50,000 deaths worldwide each year. [0003] Neisseria menin...

Claims

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Application Information

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IPC IPC(8): A61K38/02A61P31/04C07K14/22
Inventor 范玉柱
Owner BEIJING SANROAD BIOLOGICAL PROD CO LTD
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