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Pneumo-streptococcal-polysaccharide adventitia jointed vaccine and preparing method

A technology of Streptococcus pneumoniae and outer membrane protein, applied in the direction of carrier-binding antigen/hapten components, antibacterial drugs, drug combinations, etc., can solve the problems of poor immune effect of polysaccharide vaccines and uneven induction performance

Inactive Publication Date: 2007-08-29
CHANGHUI BIOLOGICAL ENG FUZHOU
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The 23-valent polysaccharide vaccine can effectively produce a protective immune response for adults, but for the elderly, infants under 2 years old, and B-cell immunodeficiency crowd, Poor immunity from polysaccharide vaccines
It has been reported that when the above-mentioned immune protein is used as a protein carrier, the induction of specific antibodies to the capsular polysaccharide of Streptococcus pneumoniae in the conjugate is different (Vaccine 2001, 19: 1159-1166; Infection And Immunity 1999, 67 (9 ): 4862-4869)

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] Example 1: Cultivation of Streptococcus pneumoniae 6B and extraction and purification of capsular polysaccharide Pn6B-PS

[0018] 1. Culture of Streptococcus pneumoniae 6B

[0019] Methods for culturing pneumococci are well known in the art (Methods of immunology and lmmunochemistry 1967, 1:52-56).

[0020] Open the ampere tube containing the freeze-dried Streptococcus pneumoniae 6B culture, add 0.5ml of normal saline, and inoculate it on the agar plate of Streptococcus pneumoniae medium containing 10% defibrated sheep blood, at 37°C and 5% CO 2 conditions for 24 hours. The culture was proved to be uncontaminated by microscopy and by agglutination with specific antiserum (6B) provided by the Danish National Blood Institute.

[0021] The above culture was inoculated into several 1-liter Erlenmeyer flasks (0.8 liters of Streptococcus pneumoniae culture medium containing peptone, glucose, and inorganic salts), and cultured at 37±0.5°C for about 12-18 hours. Use 1Mol / L s...

Embodiment 2

[0031] Example 2 Cultivation of Streptococcus pneumoniae Pn1 and extraction and purification of capsular polysaccharide Pn1-Ps

[0032] 1. Culture of Streptococcus pneumoniae Pn1

[0033] Open the ampere tube containing the freeze-dried Streptococcus pneumoniae Pn1 culture, add 0.5ml of normal saline, and inoculate it on the agar plate of Streptococcus pneumoniae medium containing 10% defibrated sheep blood, at 37°C and 5% CO 2 conditions for 24 hours. The culture was proved to be uncontaminated by microscopy and by agglutination with specific antiserum (Pn1) provided by the Danish National Blood Institute.

[0034] The above culture was inoculated into several 1-liter Erlenmeyer flasks (0.8 liters of pneumococcal culture medium containing peptone, glucose, and inorganic salts), and cultured at 37°C±0.5°C. Use 1Mol / L sodium carbonate solution to adjust the pH value of the culture medium to keep it at 6.5±0.1. Take regular samples and monitor the cell density (660nm). At th...

Embodiment 3

[0044] Example 3: Extraction, purification and immunogenicity of Streptococcus pneumoniae Pn6B outer membrane protein (Pn6B-OMPC)

[0045] The phenol-treated Streptococcus pneumoniae Pn6B culture was continuously centrifuged at low temperature to obtain supernatant and cell pellet, respectively. 300 liters of culture were centrifuged to obtain about 9000 g of cell pellet.

[0046] Extraction and purification of outer membrane proteins were performed using the following steps:

[0047] Step 1: Extraction of outer membrane proteins

[0048] Take 100 g of wet cell pellet, wash with 50 mM pH7.4 PBS buffer three times, and centrifuge at 8000 g for 15 min at 4°C. Suspend the bacterial cell sediment in PBS buffer solution containing 0.1% sodium deoxycholate at pH 7.4, and vigorously stir at 4°C for 5 hours. Filter with a 0.2um hollow fiber funnel, collect the bacterial sediment, and repeat the protein extraction for a total of 3 times. The supernatant was collected by centrifugat...

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PUM

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Abstract

The present invention relates to a streptococcus pneumoriae polysaccharide-outer membrane protein combined vaccine and its preparation method, belonging to the field of streptococcus pneumoriae vaccine. The main antigen component of said vaccine is a streptococcus pneumoriae capsular polysaccharide-outer membrane protein combined product obtained by covalently connecting the capsular polysaccharide produced by streptococcus pneumoriae with its outer membrane protein. Said capsular polysaccharide is the capsular polysaccharide of one or several kinds of streptococcus pneumoriae, its molecular weight is about 200-500 KDa, every polysaccharide molecule has about 300-700 repeating units. The outer membrane protein is the outer membrane protein of one or several kinds of streptococcus pneumoriae, its molecular weight is about 30-100 Kda. Said vaccine can be used for preventing or curing the diseases induced by streptococcus pneumoriae.

Description

technical field [0001] The invention belongs to Streptococcus pneumoniae vaccine, in particular to a novel Streptococcus pneumoniae capsular polysaccharide-outer membrane protein conjugate and a preparation method thereof. Background technique [0002] Streptococcus pneumoniae (streptococcus pneumoniae), also known as Diplococcus pneumoniae, belongs to the genus Streptococcus. Streptococcus pneumoniae can cause a variety of diseases with a wide range of prevalence, mainly causing lobar pneumonia, but also meningitis, otitis media, pleurisy, endocarditis, sepsis and other diseases. In the oral cavity and nasopharyngeal cavity of normal people, this bacterium often exists, forming a carrier state, causing diseases when the body's resistance declines, especially the elderly, immunodeficiency or immunocompromised people. [0003] Among the patients with acquired pneumonia in my country, pneumonia caused by Streptococcus pneumoniae can reach 46%-76%. Preschool children, especial...

Claims

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Application Information

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IPC IPC(8): A61K39/09A61K47/48A61P31/04A61P11/00A61K39/385
Inventor 郭养浩孟春石贤爱林海英王航叶林
Owner CHANGHUI BIOLOGICAL ENG FUZHOU
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