KC-0984-DW

MC38-mouse-PDL1-KO-PDL1 Cell Line

59426

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Background of MC38-mouse-PDL1-KO-PDL1 Cell Line

PD-L1, also called human programed cell death ligand 1, is a transmembrane protein that play a major role in suppressing the immune system during particular events such as pregnancy, tissue allografts, autoimmune disease, virus infection and cancer. PD-L1 binds to its receptor PD-1 on activated T cells, B cells, and myeloid cells, to modulate activation or inhibition. Upregulation of PD-L1 can allow cancer cell to evade the host immune system.

Specifications

Catalog Number	KC-0984-DW
Cell Line Name	MC38-mouse-PDL1-KO-PDL1 Cell Line
NCBI/UniProt Accession Number	NM_014143.4
Clone Number	NA
Host Cell Line	MC38
Description	Stable MC38-mouse-PDL1-KO cell line expressing exogenous PDL1 gene.
Quantity	One vial of frozen cells (≥2-10⁶/vial)
Stability	Stable in culture over a minimum of 10 passages
Application	Drug screening and biological assays
Freezing Medium	70% DMEM+20% FBS+10% DMSO
Propagation Medium	DMEM+15%FBS+0.01mM NEAA+100μg/mL Hygromycin B
Selection Marker	Hygromycin B
Morphology	Epithelial
Subculture	Split saturated culture 1:4-1:10 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL
Incubation	37 °C with 5% CO₂
Storage	Liquid nitrogen immediately upon receiving
Doubling Time	Approximately 30 hours
Mycoplasma Status	Negative

Cell Line Generation

MC38-mouse-PDL1-KO-PDL1 cell line was generated using lentivirus expressing PDL1 sequence.

Characterization

Figure 1. Characterization of mouse-PDL1 knockout and PDL1 over-expression in the MC38-mouse-PDL1-KO-PDL1 stable clone using FACS.

Figure 2:Validation of in vivo tumorigenicity of MC38-mouse-PDL1-KO cells stably expressing PDL1 via subcutaneous implantation in C57BL/6J mice, with tumor growth monitored by measuring volume. (V=0.5×length×width²)

Figure 3. Characterization of PDL1 over-expression in the MC38-mouse-PDL1-KO-PDL1 stable clone using PCR.

Cell Resuscitation

Prewarm culture medium (DMEM supplemented with 15% FBS, 0.01mM NEAA+100μg/mL Hygromycin B)in a 37°C water bath.
Thaw the frozen vial in a 37°C water bath for 1-2 minutes.
Transfer the vial into biosafety cabinet, and wipe the surface with 70% ethanol.
Unscrew the top of the vial and transfer the cell suspension gently into a sterile centrifuge tube containing 9.0mL complete culture medium.
Spin at ~ 125 × g for 5-7 minutes at room temperature, and discard the supernatant without disturbing the pellet.
Resuspend cell pellet with the appropriate volume of complete medium and transfer the cell suspension into a T25 culture flask.
Incubate the flask at 37°C, 5% CO₂ incubator.
Split saturated culture 1:4-1:10 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

Prepare the freezing medium (70% DMEM + 20% FBS + 10% DMSO) fresh immediately before use.
Keep the freezing medium on ice and label cryovials.
Transfer cells to a sterile, conical centrifuge tube, and count the cells.
Centrifuge the cells at 250×g for 5 minutes at room temperature and carefully aspirate off the medium.
Resuspend the cells at a density of at least 3×10⁶ cells/mL in chilled freezing medium.
Aliquot 1 mL of the cell suspension into each cryovial.
Freeze cells in the CoolCell freezing container overnight in a -80°C freezer.
Transfer vials to liquid nitrogen for long-term storage.

References

Akinleye, Akintunde, Yamei Chen, Nikhil Mukhi, Yongping Song, and Delong Liu. 2013. “Ibrutinib and Novel BTK Inhibitors in Clinical Development.” Journal of Hematology & Oncology 6 (1). Journal of Hematology & Oncology: 1–1. doi:10.1186/1756-8722-6-59.
Cheng, S, A Guo, P Lu, J Ma, M Coleman, and Y L Wang. 2014. “Functional Characterization of BTKC481S Mutation That Confers Ibrutinib Resistance: Exploration of Alternative Kinase Inhibitors,” September. Nature Publishing Group, 1–25. doi:10.1038/leu.2014.263.

Use License Agreement

Research Use Only.

Not for use in diagnostic procedures or therapeutic applications.

Redistribution of the cell line or its derivatives is prohibited without prior written permission from Kyinno Biotechnology.