KC-5232

293T-mouse-BCMA Cell Line

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Background of 293T-mouse-BCMA Cell Line

B cell maturation protein (BCMA), also named CD269, is a membrane of the TNF receptor super family (TNFRSF17) and mainly expressed in mature B-lymphocytes, and play an important role in B cell development and autoimmune response. One of the biggest features of BCMA targeted therapy is that it only focuses on one disease, because BCMA is almost not expressed in other tissue cells except on the surface of mature B lymphocytes and plasma cells, but it is highly expressed in all MM cells, and plays a key role in the survival, proliferation, metastasis and drug resistance of MM cells through mediating the downstream signaling pathway. So BCMA is an ideal antigenic target for the treatment of MM.

Specifications

Catalog Number	KC-5232
Cell Line Name	293T-mouse-BCMA Cell Line
Clone Number	5-3#
Host Cell Line	293T
Description	Stable 293T clone expressing exogenous mouse BCMA gene
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% DMEM + 20% FBS + 10% DMSO
Propagation Medium	DMEM + 10% FBS + 1μg/mL Puromycin
Selection Marker	Puromycin
Morphology	Epithelial
Subculture	Split saturated culture 1:4-1:8 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

293T-mouse-BCMA cell line was generated using a lentiviral vector expressing the mouse BCMA sequence.

Characterization

Figure 1: Characterization of mouse BCMA overexpression in the 293T stable clone using FACS(Kyinno, Cat#KB-1514).

Figure 2: Characterization of mouse BCMA in the 293T stable clone using PCR sequencing.

Cell Resuscitation

1. Prewarm culture medium (DMEM supplemented with 10% FBS and 1μg/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 1-3 × 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. Gross, J. A. et al. TACI and BCMA are receptors for a TNF homologue implicated in B-cell autoimmune disease. Nature 404, 995ÿ999 (2000).
2. Yu B, Jiang T, Liu D. BCMA-targeted immunotherapy for multiple myeloma. J Hematol Oncol. 2020 Sep 17;13(1):125. doi: 10.1186/s13045-020-00962-7. PMID: 32943087; PMCID: PMC7499842.
3. Shah N, Chari A, Scott E, Mezzi K, Usmani SZ. B-cell maturation antigen (BCMA) in multiple myeloma: rationale for targeting and current therapeutic approaches. Leukemia. 2020 Apr;34(4):985-1005. doi: 10.1038/s41375-020-0734-z. Epub 2020 Feb 13. PMID: 32055000; PMCID: PMC7214244.
4. Sun F, Cheng Y, Wanchai V, Guo W, Mery D, Xu H, Gai D, Siegel E, Bailey C, Ashby C, Al Hadidi S, Schinke C, Thanendrarajan S, Ma Y, Yi Q, Orlowski RZ, Zangari M, van Rhee F, Janz S, Bishop G, Tricot G, Shaughnessy JD Jr, Zhan F. Bispecific BCMA/CD24 CAR-T cells control multiple myeloma growth. Nat Commun.2024 Jan 19;15(1):615. doi: 10.1038/s41467-024-44873-4. PMID: 38242888; PMCID:PMC10798961.