KC-1755

H22-mouse-CD274-KO-3A3-Cell-Line

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Background of H22-mouse-CD274-KO-3A3-Cell-Line

CD27, also named as TNFRSF7, is a transmembrane protein belonging to the TNF receptor superfamily, and not only functioned as a co-stimulatory immune checkpoint molecule in T cell activation, but also play a critical role in regulating B-cell activation and immunoglobulin synthesis after binding with its ligand, CD70. CD27 is also promising immunotherapy target for cancer treatment, its monoclonal antibody Varlilumab are in phase I for solid tumor and hematologic malignancies.

Specifications

Catalog Number	KC-1755
Cell Line Name	H22-mouse-CD274-KO-3A3-Cell-Line
Host Cell Line	Mouse H22 cell line
Description	H22 cell line stable clone with mouse CD274 gene knockout
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% RPMI1640+20% FBS+10% DMSO
Propagation Medium	RPMI1640+10%FBS
Morphology	Round
Subculture	Split saturated culture 1:4-1:5 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
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

H22 mouse-CD274 KO cell line was generated using the CRISPR method.

Characterization

Cell Resuscitation

Prewarm culture medium (RPMI-1640 supplemented with 10% FBS)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 1:4-1:5 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

Lens SM, de Jong R, Hintzen RQ, Koopman G, van Lier RA, van Oers RH (Jun 1995). "CD27-CD70 interaction: unravelling its implication in normal and neoplastic B-cell growth". Leukemia & Lymphoma. 18 (1ÿ2): 51ÿ9.

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.