组织型纤溶酶原激活剂(t-PA)突变体工程细胞株生物学特性分析  

THE ANALYSIS OF BIOLOGICAL CHARACTERISTICS FOR THE TISSUE-PLASMINOGEN ACTIVATOR (t-PA) MUTANT ENGINEERED CELL LINE

作　　者：陈琳[1] 徐秀英[1] 刘士辉[1] 朱恒奇[1] 黄培堂[1] 黄翠芬[1] 

机构地区：[1]军事医学科学院生物工程研究所,北京100071

出　　处：《实验生物学报》1998年第2期111-116,共6页Acta Biologiae Experimentalis Sinica

摘　　要：t-PA突变体工程细胞株FSGGI48形态与其亲代细胞CHO-dhfr相似呈多角形,类似上皮细胞。在MTX加压至5×10^(-6)mol/L时,少数细胞形态略变瘦长,但仍呈多角形,因此该工程细胞株形态正常。抗体中和抑制试验及纤维蛋白板80℃加热试验显示,FSGGI48细胞表达产物与st-PA的活性均能被抗t-PA抗体所抑制,而胰酶的活性则不被抑制,且二者在80℃加热的纤维蛋白板上均不产生溶解圈,胰酶则产生溶解圈,由此可知,该细胞株表达产物为特异的rt-PA产品。细胞无血清培养上清经FA-PA检测,亚克隆株表达水平为4000IU/10~6细胞/24h。分别测定冻存3个月后复苏和体外传代3个月以上细胞的表达水平,结果显示部分亚克隆细胞株表达水平下降,多数仍为3000-4000IU/10~6细胞/24h,说明细胞株稳定性好。裸鼠试验表明,该工程株活细胞、细胞DNA、纯化的细胞表达产物均无致瘤性。支原体检查结果为阴性。染色体分析显示,FSGGI48细胞与其亲代细胞(CHO-dhfr细胞)染色体数目相同均为20条,但均有不同程度不同类型的畸变。CHO-dhfr细胞畸变率为6%。该工程细胞的畸变率为15%-18%,在允许范围内。结果证明,t-PA突变体细胞株FSGGI48为性能优良的工程细胞株。The morphology of t-PA mutant engineered cell line FSGGI48 is similar to that of CHO-dhfr cell in polygon. When aminopterin (MTX) concentration is 5×10-6 mol/L, a part of cells become slim, but they are still in ploygon. Therefore , the morphology of FSGGI48 cell line is normal. The expression level is 4000IU/106cells/24h in serum-free medium. The cell line is bequeathed for three months in vitro and revived after freezing three months, the greater part of cells are stable. The expression level is 3000 - 4000IU/106celIs/24h. The assay of tumorigene-sis shows that the cells, cellular DNA and purified products don't form tumors in nude mice. The cell line hasn't been contaminated by my-coplasma. The results of chromosomal analysis show that the chromosomal number of CHO-dhfr cell is 20 and the abnormal rate is 6%. FSGGI48 cell line is same as CHO-dhfr- cell in chromosomal number and the abnormal rate is 15%-18%, it is normal in scope. Therefore, the engineered cell line FSGGI48 is excellent.

关 键 词：T-PA 突变体 工程细胞株 生物学特性 

分 类 号：Q78[生物学—分子生物学]