TF, also known as CD142, is a type I transmembrane glycoprotein, which is expressed on the surface of a variety of cells that are physically separated from the circulating blood which include smooth muscle cells, fibroblasts, keratinocytes, glomerular epithelial cells (cytoplasmic inclusions), astrocytes, myocardium, liver stromal cells, pancreas cells, and is also expressed on activated monocytes and stimulated endothelial cells. TF is a high-affinity receptor for coagulation factor VII and initiates the extrinsic pathway of blood coagulation. TF also plays an important role in a variety of diseases such as sepsis, atherosclerosis, and cancer.
Cell Line Name:
CHO-K1 TF Cell Line
Host Cell Line:
Chinese hamster ovary CHO-K1 cell line
Stable CHO-K1 clone expressing exogenous human TF
One vial of frozen cells (5X106 per vial)
Stable in culture over a minimum of 10 passages
Drug screening and biological assays
70% RPMI1640+20% FBS+10% DMSO
Split saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-2 × 105 cells/mL
37 °C with 5% CO2
Liquid nitrogen immediately upon receiving
Approximately 24 hours
Cell Line Generation
CHOK1 human TF cell line was generated using a lentiviral vector expressing the human TF sequence.
Characterization using PCR sequencing and WB
1. Prewarm culture medium (RPMI1640 + 10% FBS + 10μg/mL Puromycin) in a 37°C water bath.
2. Thaw the frozen vial in a 37°C water bath for 1-2 minutes.
3. Transfer the vial into biosafety cabinet, and wipe the surface with 70% ethanol.
4. Unscrew the top of the vial and transfer the cell suspension gently into a sterile centrifuge tube containing 9.0mL complete culture medium.
5. Spin at ~ 125 × g for 5-7 minutes at room temperature, and discard the supernatant without disturbing the pellet.
6. Resuspend cell pellet with the appropriate volume of complete medium and transfer the cell suspension into a T25 culture flask.
7. Incubate the flask at 37°C, 5% CO2 incubator.
8. Split saturated culture 1:4-1:8 every 2-3 days; seed out at about 1-2 × 105 cells/mL.
1. Prepare the freezing medium (70% RPMI-1640 + 20% FBS + 10% DMSO) fresh immediately before use.
2. Keep the freezing medium on ice and label cryovials.
3. Transfer cells to a sterile, conical centrifuge tube, and count the cells.
4. Centrifuge the cells at 250×g for 5 minutes at room temperature and carefully aspirate off the medium.
5. Resuspend the cells at a density of at least 3×106 cells/mL in chilled freezing medium.
6. Aliquot 1 mL of the cell suspension into each cryovial.
7. Freeze cells in the CoolCell freezing container overnight in a -80°C freezer.
8. Transfer vials to liquid nitrogen for long-term storage
1. "Entrez Gene: F3 coagulation factor III (thromboplastin, tissue factor)".
2. Muller YA, Ultsch MH, de Vos AM (February 1996). "The crystal structure of the extracellular domain of human tissue factor refined to 1.7 A resolution". Journal of Molecular Biology. 256 (1): 144 – 59. doi:10.1006/jmbi.1996.0073. PMID 8609606.
3. Zhang E, St Charles R, Tulinsky A (February 1999). "Structure of extracellular tissue factor complexed with factor VIIa inhibited with a BPTI mutant". Journal of Molecular Biology. 285 (5): 2089 – 104. doi:10.1006/jmbi.1998.2452. PMID 9925787.
4. Tissue factor in hematological malignancies. Versteeg, H.H. et al. (2003) Carcinogenesis 24:1009.
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