KC-5751

MC38-B7H3-GFP-Luc2-High Cell Line

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Background of MC38-B7H3-GFP-Luc2-High Cell Line

The immune checkpoint molecule B7-H3, also known as CD276, is a membrane protein member of the B7-CD28 family of immunomodulatory proteins, a type I membrane protein that is sequence-similar to the extracellular domain of programmed death receptor-1 (PD-L1). In normal human tissues, the expression level of B7-H3 is low, but in a variety of tumor cancers, B7-H3 is abnormally high, which plays an important role in the development of tumors, immune escape and other processes, and is associated with the poor prognosis of tumors. At present, there are a total of 109 biologics developed for B7-H3 targets, but only 33 are in the clinical stage, of which ADC and CAR-T therapy are the main ones.

Specifications

Catalog Number	KC-5751
Cell Line Name	MC38-B7H3-GFP-Luc2-High Cell Line
Clone Number	4#
Host Cell Line	MC38
Description	Stable MC38 cell line expressing exogenous B7H3 GFP Luc2 gene in high level
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 + 10% FBS
Selection Marker	NA
Morphology	Epithelial
Subculture	Split saturated culture 1:4~1:8 every 2~3 days; seed out at about 1-2 × 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 B7H3 GFP Luc2 Cell Line was generated using a lentiviral vector expressing the B7H3 GFP Luc2 sequence.

Characterization

Figure 1: Characterization of B7H3 GFP Luc2 overexpression in the MC38 B7H3 GFP Luc2 stable clone using FACS.

Figure 2: Characterization of luciferase expression in MC38 B7H3 GFP Luc2 stable clone using the Bright-Lite Luciferase Detection System.

Figure 3: Characterization of B7H3 GFP Luc2 overexpression in the MC38 B7H3 GFP Luc2 stable clone using PCR sequence.

Cell Resuscitation

1. Prewarm culture medium (DMEM supplemented with 10% FBS)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.

Cell Freezing

1. Prepare the freezing medium (70% DMEM + 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.

References

1.Dong P, Xiong Y, Yue J, Hanley SJB, Watari H. B7H3 As a Promoter of Metastasis and Promising Therapeutic Target. Front Oncol. 2018 Jul 6;8:264.

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.