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地鼠肾细胞狂犬病疫苗原液经100 kD 膜浓缩 30 倍,分别选用(1)DEAE Sepharose CL-6B离子交换层析法;(2)Sephacry1 S-200 HR 分子筛选层析法;(3)二次蔗糖等密度区带离心法对其进行纯化。用此三种方法各试制3 批精制疫苗,结果表明,经DEAE Sepharose CL-6B离子交换层析纯化后疫苗总蛋白含量减少99% 以上,抗原比活性提高159 倍,抗原回收率达50% ,纯化疫苗以NIH 法效力测定平均为5.4 IU/2m l;经Sephacry1 S-200HR 分子筛层析纯化后疫苗总蛋白含量减少 98% 以上,抗原比活性提高41 倍,抗原回收率达63% ,纯化疫苗效力平均为6.25 IU/2m l;经一次蔗糖等密度区带离心法纯化后疫苗总蛋白含量减少98% 以上,抗原比活性提高321 倍,抗原回收率达43% ,纯化疫苗效力平均为6.18 IU/2m l,三种纯化疫苗均符合W HO 规程要求。
The hamster kidney cell rabies vaccine stock solution was concentrated 30-fold by 100 kD membrane, respectively, (1) DEAE Sepharose CL-6B ion exchange chromatography; (2) Sephacryl S-200 HR molecular sieve chromatography; (3) Sucrose and other isolates were purified by centrifugation. Three batches of the purified vaccine were tested by these three methods. The results showed that the total protein content of the vaccine purified by DEAE Sepharose CL-6B ion exchange chromatography was reduced by more than 99%, the specific activity of the antigen was increased by 159 times and the antigen recovery rate was 50% The purified vaccine had an average of 5.4 IU / 2 ml measured by NIH method. The total protein content of the vaccine purified by Sephacryl S-200HR chromatography was reduced by more than 98%, the specific activity of the antigen was increased by 41 times and the antigen recovery rate was 63% The average potency of the purified vaccine was 6.25 IU / 2m l. The total protein content of the vaccine was reduced by more than 98% after a single centrifugation in sucrose density zone, the specific activity of the antigen was increased by 321 times and the antigen recovery rate was 43% With an average of 6.18 IU / 2m l. All three purified vaccines met the W HO protocol.