Peiwei G, Zhifen D, Zhongquan W
National Vaccine and Serum Institute, Beijing, China.
Dev Biol Stand. 1989;70:223-6.
Two continuous cell lines, lymphoblastoid cell (Namalwa) and BHK-21, were studied for production of interferon and Japanese Encephalitis vaccine. Lymphoblastoid cells (Namalwa) were induced with 100 HAU/ml of Sendai virus while the cell density was 10(7) cells/ml. A titre of Ig 4.0-5.01 U/ml interferon was obtained at 16-18 hours after induction. A semi-continuous production technique was established in a fermentor. Thirty days of cell cultivation and induction was carried out continuously with a regular production of interferon. The crude lymphoblastoid interferon was purified with precipitation with trichloro-acetic acid, buffer solution treatment and blue-sepharose chromatography. The specific activity reached 10(6)-10(7) I.U/mg. No residual DNA was detected in the purified interferon. Four malignant tumour patients were treated with lymphoblastoid interferon. The preliminary result of the clinical trial was encouraging. BHK-21 C13 cell was tested for the karyological characteristics and its sensitivity to Japanese Encephalitis (JE) virus. No karyological change was observed after 52 passages. After infection of the monolayer cell culture with JE virus, a typical cytopathic effect (CPE) was observed. The virus titre in the maintenance medium rose gradually while the CPE developed and more than log TCID50 7.0/0.1 ml could be obtained. After inactivation with 1:2000 formalin, a vaccine of high potency was successfully produced. It is expected that a suspension culture technique using a fermentor will give a higher titre of virus. A large scale production of inactivated vaccine will be succeeded when the characterization of BHK-21 C13 cell line and purification procedure are standardized.
对两种连续细胞系，即淋巴母细胞样细胞（Namalwa）和BHK - 21，进行了干扰素和日本脑炎疫苗生产的研究。用100 HAU/ml仙台病毒诱导淋巴母细胞样细胞（Namalwa），此时细胞密度为10⁷个细胞/ml。诱导后16 - 18小时获得了4.0 - 5.01 U/ml干扰素的滴度。在发酵罐中建立了半连续生产技术。连续进行30天的细胞培养和诱导，可定期生产干扰素。粗制的淋巴母细胞样干扰素通过三氯乙酸沉淀、缓冲液处理和蓝琼脂糖层析进行纯化。比活性达到10⁶ - 10⁷ I.U/mg。纯化后的干扰素中未检测到残留DNA。用淋巴母细胞样干扰素治疗了4例恶性肿瘤患者。临床试验的初步结果令人鼓舞。对BHK - 21 C13细胞进行了染色体特征及其对日本脑炎（JE）病毒敏感性的检测。传代52次后未观察到染色体变化。用JE病毒感染单层细胞培养物后，观察到典型的细胞病变效应（CPE）。维持培养基中的病毒滴度随着CPE的发展而逐渐升高，可获得超过log TCID50 7.0/0.1 ml的病毒滴度。用1:2000福尔马林灭活后，成功生产出高效疫苗。预计使用发酵罐的悬浮培养技术将产生更高滴度的病毒。当BHK - 21 C13细胞系的特性和纯化程序标准化后，灭活疫苗的大规模生产将取得成功。
