KC-3382

CHOK1-cyno-CD20-Cell-Line

26979

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Background of CHOK1-cyno-CD20-Cell-Line

CD20, This gene encodes a member of the membrane-spanning 4A gene family. Members of this nascent protein family are characterized by common structural features and similar intron/exon splice boundaries and display unique expression patterns among hematopoietic cells and nonlymphoid tissues. This gene encodes a B-lymphocyte surface molecule which plays a role in the development and differentiation of B-cells into plasma cells. This family member is localized to 11q12, among a cluster of family members. Alternative splicing of this gene results in two transcript variants which encode the same protein. CD20 is highly expressed in several types of B-cell lymphoma and is an intuitive target for chimeric antigen receptor (CAR) T-cell therapy.

Specifications

Catalog Number	KC-3382
Cell Line Name	CHOK1-cyno-CD20-Cell-Line
Host Cell Line	CHOK1
Description	Stable CHOK1 clone expressing exogenous cyno-CD20 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% RPMI 1640 + 20% FBS + 10% DMSO
Propagation Medium	RPMI 1640 + 10% FBS + 10μg/mL Puromycin
Selection Marker	Puromycin
Morphology	Epithelial
Subculture	Split saturated culture 1:6-1:8 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 24 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

CHOK1-cyno-CD20-cell-line was generated using a lentiviral vector expressing the cyno-CD20 sequence.

Characterization

Figure 1: Characterization of cyno-CD20 overexpression in the CHOK1-cyno-CD20 stable clone using FACS.

Figure 2: Characterization of cyno-CD20 overexpression in the CHOK1 stable clone using FACS.

Cell Resuscitation

Prewarm culture medium (RPMI 1640 supplemented with 10% FBS and 10μg/mL puromycin)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 1:6-1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

Prepare the freezing medium (70% RPMI 1640 + 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

Scheller L, Hudecek M. Engineering CD20 CARs with a Twist. Cancer Immunol Res. 2023 Feb 3;11(2):142-143. doi: 10.1158/2326-6066.CIR-22-0919. PMID: 36633575.
Kim JR, Lee D, Kim Y, Kim JY. CD20/TNFR1 dual-targeting antibody enhances lysosome rupture-mediated cell death in B cell lymphoma. Cancer Immunol Immunother. 2023 Jun;72(6):1567-1580. doi: 10.1007/s00262-022-03344-9. Epub 2022 Dec 19. PMID: 36534148.