KC-5746

CT26-B7H3-GFP-Luc2-High Cell Line

59568

Home » CT26-B7H3-GFP-Luc2-High Cell Line

Background of CT26-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-5746
Cell Line Name	CT26-B7H3-GFP-Luc2-High Cell Line
Clone Number	6#
Host Cell Line	CT26
Description	Stable CT26 cell line expressing exogenous B7H3 GFP Luc2 gene in high level
Quantity	Two vials 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% RPMI1640 + 20% FBS + 10% DMSO
Propagation Medium	RPMI1640 + 10% FBS
Selection Marker	NA
Morphology	Epithelial
Subculture	Split saturated culture 1:4~1:6 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

CT26 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 CT26 B7H3 GFP Luc2 stable clone using FACS.

Figure 2: Characterization of luciferase expression in CT26 B7H3 GFP Luc2 stable clone using the Bright-Lite Luciferase Detection System.

Figure 3: Characterization of B7H3 GFP Luc2 overexpression in the CT26 B7H3 GFP Luc2 stable clone using PCR sequence.

Cell Resuscitation

1. Prewarm culture medium (RPMI1640 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 1:4~1:6 every 2~3 days; seed out at about 1-2 × 10⁵ cells/mL.

Cell Freezing

1. Prepare the freezing medium (70% RPMI1640 + 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.