Establishment and characterization of new human cell lines for recombinant therapeutic protein production
© Inoue et al. 2015
Published: 14 December 2015
Recombinant therapeutic proteins are increasingly requested with advances in tissue engineering using stem cells. Human cell line is an attractive host for the production of such glycoprotein, but there are few reports on human cells for a commercial production . In this study, we established new human lymphoid cell lines from peripheral blood mononuclear cells (PBMCs) by treatment with phorbol 12-myristate 13-acetate (PMA) under a non-GMP condition, and characterized them by gene and protein expression analyses.
The human PBMCs (2 × 106 cells/ml) were cultured in 24 well plates in 12.5%FBS-ERDF medium supplemented with 10 ng/ml of IL-4 and/or IL-6 and 1 µg/ml of PMA for three months. The medium was changed every two or three days.
The gene expression of telomerase reverse transcriptase (TERT), a marker of immortalization, was examined by RT-PCR. The immunoglobulin (Ig) isotype was confirmed by ELISA. The CD markers on cell surface were detected by flow cytometry.
Results and Discussion
Summary of cell establishment and characterization.
Culture medium for transformation
Culture medium for maintenance
Possible period of subculture
Over 1 year
Over 1 year
Over 2 years
Mean doubling time
B cell marker (CD19)
This work was supported by JSPS KAKENHI Grant Number 26460167.
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