STABLE CELL-LINE GENERATION

Trican Biotech. provides stable mammalian expression cell line construction services using proprietary vectors with selectable markers to achieve high productivity in serum-free/chemically-defined medium. Through single-cell cloning from resistant populations, the selected production line is suitable for research use and for industrial manufacturing of biologics, antibodies, and engineered proteins.

Features

1. Robust and high-productivity cell line development

2. Quick turnaround time

3. Suitable for antibody, biologic, or engineered protein

4. Capability to develop clones at 0.5~1g/L prior to culture process optimization

5. Cell line characterizations including biosafety, expression stability test, etc

Stable cell line construction flow chart

Stable Erythropoietin expression CHO cell line construction and characterization

Cells were grown in CD OptiCHO™ Medium (Invitrogen, USA) for three passages prior to the test. Cells were seeded at 3 × 105/ml in 30 mL working volume in a 125 mL shake flask. Cultures were incubated at 35°C, 8% CO2, and shaken at 125 rpm. Viable cell density determined using LUNA-IITM Automated Cell Counter (Logos Biosystems, Korea). Amount of Erythropoietin in culture was determined by ELISA using EPREX® 2000 (Janssen-Cilag Ltd., UK) as standard. The calculated Erythropoietin concentration is equivalent to 660mg/L at Day 12 under shaker flask in the test culture.

Detection of recombinant Erythropoietin expressed

by a stable CHO expression line

Recombinant Erythropoietin in culture supernatants were detected using Western blot analysis with EPREX® 2000 (Janssen-Cilag Ltd., UK) as control. The data demonstrated that the expressed recombinant Erythropoietin had the similar pattern as that of EPREX® 2000 (Epoetin alpha).

Lane 1: 40ul of culture supernatant after cultured 10 days

Lane 2: 40ul of culture supernatant after cultured 11 days

Lane 3: 1ug Epoetin alfa (Mw. is 30 to 40kD)

M: Protein molecular weight markers as indicated

Cells were grown in CD OptiCHO™ Medium (Invitrogen, USA) for three passages prior to the test. Cells were seeded at 3 × 105/ml in 30 mL working volume in a 125 mL shake flask. Cultures were incubated at 35°C, 8% CO2, and shaken at 125 rpm. Viable cell density determined using LUNA-IITM Automated Cell Counter (Logos Biosystems, Korea). Amount of Erythropoietin in culture was determined by ELISA using EPREX® 2000 (Janssen-Cilag Ltd., UK) as standard. The calculated Erythropoietin concentration is equivalent to 660mg/L at Day 12 under shaker flask in the test culture.