Objective To study the establishment of cell engineering laboratory of blood cells polymorphism through establishing immortalized lymphoblastoid cell lines( LCLs ) from the rare antigens of blood cells. Methods Immortalized cell lines of lymphocytes were established from rare antigens of red cells, known antigen type of neutrophils, and the rare antigens of platelets as well as the CD36 deficiency platelets by ￡pstein-barr( EB ) virus transformation, in which mycoplasma test was performed with culture assay and polymerase chain reaction( PCR ). Genetic stability of cell line was determined by the analysis of chromosome karotype. Results We have successfully estabolished 211 immortalized cell lines,including 64 LCLs of the rare antigens of red cells,8 LCLs of the known antigen type of neutrophils, 15 LCLs of the rare antigens of platelets and 124 LCLs of the antigens of CD36 deficiency platelet. The total transformation rate was 91% ,the subculture and culture recovery rate was 100%, which provided eternal study materials for the cell engineering lab. No gene mutation and mycoplasma contamination appeared. No chromosomal mutation occurred at 20th passage. Conclusion The techniques of subculturing, cryogenic preservation, culture recovery and mycoplasma contamination prevention could meet the requirement of establishment of the cell line panel, which provides the scientific references for the establishment of cell engineering laboratory of blood cells lymorphism.%目的 建立稀有型血细胞永生化细胞株,探讨血细胞多态性细胞工程实验室建立的方法.方法 采用EB病毒转化技术,把携带稀有血型抗原的红细胞、已知抗原特异性的粒细胞,罕见特异性抗原的血小板及CD36缺失型的血小板的外周血淋巴细胞转化成永生化淋巴母细胞株,采用培养法和PCR法对细胞株进行支原体检测,染色体核型分析细胞株的稳定性.结果 共建立稀有型血细胞LCLs 211株,其中稀有红细胞血型抗原LCLs 64株、已知抗原特异性的粒细胞抗原LCLs 8株、罕见特异性抗原的血小板LCLs 15株及CD36缺失型血小板LCLs 124株,总转化成功率为91%;所有的细胞株传代、冻存和复苏均成活,无基因突变、无支原体污染.经20代传代后未发现染色体突变.结论 传代培养、细胞冻存、复苏和支原体污染防范等一整套技术过程基本满足建库要求,为血细胞多态性细胞工程实验室的建立提供科学依据.
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