HBsAg, technology for preparing HBsAg through expression by virtue of recombinant saccharymyces cerevisiae, separation and purification technology for HBsAg as well as hepatitis B vaccine

Abstract

The invention discloses a technology for preparing HBsAg through expression by virtue of recombinant saccharymyces cerevisiae and a separation and purification technology of the HBsAg. The separation and purification technology of the HBsAg comprises the following steps: breaking cells, namely feeding cerevisiae cells extracted after fermentation into a high pressure homogenizer and breaking the cerevisiae cells at a pressure of 15000+ / -1500PSIG; heating feed liquid after breaking, namely heating the feed liquid to 50-80 DEG C and then cooling the feed liquid to 2-8 DEG C; and purifying, namely purifying the feed liquid and preparing HBsAg raw liquid. The invention also discloses a corresponding hepatitis B vaccine and HBsAg. The technology for preparing the HBsAg through expression by virtue of the recombinant saccharymyces cerevisiae has the advantages that protease can be inactivated without adding a protease inhibitor before the cells are broken, so that secondary pollution caused by the protease inhibitor is avoided; meanwhile, the yield of the raw liquid obtained through purification subsequently can be obviously increased, and quality indexes such as 'specific activity' and 'Sub-p24' of the raw liquid are greatly improved, so that the technology for preparing the HBsAg through expression by virtue of the recombinant saccharymyces cerevisiae has high economic benefit and social benefit.

Description

technical field [0001] The invention relates to a preparation technology of recombinant gene hepatitis B vaccine, in particular to a preparation technology of recombinant Saccharomyces cerevisiae expressing hepatitis B surface antigen (HBsAg), its separation and purification technology, HBsAg and hepatitis B vaccine. Background technique [0002] my country is a high-prevalence area of ​​hepatitis B, with an average of more than 500,000 cases reported each year, and hepatitis B vaccination is the most effective, economical and safest way to prevent hepatitis B. In 1992, my country incorporated the hepatitis B vaccine into the planned immunization management. Since 2009, the country has expanded the scope of free hepatitis B vaccination to children under the age of 15, as well as medical staff, college students and active military personnel. The demand is getting bigger and bigger, and the coverage is getting wider and wider. The implementation of the new version of GMP in 20...

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Problems solved by technology

[0005] Due to the short half-life of protease inhibitors (about 30 minutes), and in large-scale production, the time required for cell disruption is longer (60-80 minutes), which often leads to the late stage of the disruption process, and the effect of protease inhibitors has declined or disappeared. , the target substance HBsAg released by crushing is decomposed to a certain extent by protease, thus increasing the difficulty of subsequent purification, resulting in the impact on the yield and quality of the purified stock solution; in addition, because the added protease inhibitor enters the buffer system, it also brings Here comes the problem of secondary pollution

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