人尿激酶原工程细胞株 C L-11 G 细胞外源因子的检测

The detection of exogenous factors in human pro urokinase engineering cell line CL 11G

目的：对工程细胞株的要求之一是不应被外源因子污染。因此有必要对表达人尿激酶原的工程细胞株 Ｃ Ｌ１１ Ｇ 进行外源因子的检测。方法：用普通肉汤、改良马丁和硫乙醇酸盐三种培养基对 Ｃ Ｌ１１ Ｇ 细胞进行培养，检查细菌和霉菌；用培养法、 Ｄ Ｎ Ａ 荧光染色法、单克隆抗体免疫荧光法及电镜检查支原体；用 Ｃ Ｌ１１ Ｇ 细胞悬液分别接种乳小鼠、成年小鼠、豚鼠、家兔、鸡胚及 ５ 种细胞，检查有无内外源性病毒污染。结果： Ｃ Ｌ１１ Ｇ 细胞经上述几种方法检测均未发现有细菌、霉菌及支原体污染。接种的动物全部存活，无异常，５ 种细胞接种 Ｃ Ｌ１１ Ｇ 细胞后细胞形态正常，无病变，红细胞吸附试验均为阴性。结论：人尿激酶原工程细胞株 Ｃ Ｌ１１ Ｇ 细胞经多方检查，未发现细菌、霉菌、支原体及内外源性病毒污染，从细胞水平上讲是安全的，可以作为生产重组人尿激酶原溶栓药物的工程细胞株。

Objective: Cell line without contamination by exogenous factors is one of the requirements for engineering cell line, so it is necessary to examine whether the CL 11G cell line was contaminated by exogenous factors or not. Methods: The contamination of bacteria and mold was examined by incubating the CL 11G cell in three different media, the contamination of mycoplasma by means of culturing, DNA fluorescence staining, monoclonal antibody immunofluorescence and electron microscopy, and the virus contamination by inoculating the CL 11G cell suspension into different species of animals, chicken embryo and five cell lines. Results: No contamination of the CL 11G cell line by any exogenous factors such as bacteria, mold, mycoplasma or virus was found by all above tests. All inoculated animals were alive healthily. The five inoculated cell lines grew well and their morphology was normal without any pathological changes. The erythrocyte absorption tests were negative. Conclusion: The CL 11G cell line was not contaminated by any exogenous factors and can be used to produce recombinant pro urokinase safely.