KC-3785

K562-41BBL-mem-IL21 Cell Line

34772

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Background of K562-41BBL-mem-IL21 Cell Line

The protein encoded by this gene is a cytokine that belongs to the tumor necrosis factor (TNF) ligand family. This transmembrane cytokine is a bidirectional signal transducer that acts as a ligand for TNFRSF9/4-1BB, which is a costimulatory receptor molecule in T lymphocytes. This cytokine and its receptor are involved in the antigen presentation process and in the generation of cytotoxic T cells. The receptor TNFRSF9/4-1BB is absent from resting T lymphocytes but rapidly expressed upon antigenic stimulation. The ligand encoded by this gene, TNFSF9/4-1BBL, has been shown to reactivate anergic T lymphocytes in addition to promoting T lymphocyte proliferation. This cytokine has also been shown to be required for the optimal CD8 responses in CD8 T cells. This cytokine is expressed in carcinoma cell lines, and is thought to be involved in T cell-tumor cell interaction.
IL21 gene encodes a member of the common-gamma chain family of cytokines with immunoregulatory activity. The encoded protein plays a role in both the innate and adaptive immune responses by inducing the differentiation, proliferation and activity of multiple target cells including macrophages, natural killer cells, B cells and cytotoxic T cells. Dysregulation of this gene plays a role in multiple immune-mediated diseases including lupus, psoriasis and chronic inflammatory diseases. Alternatively spliced transcript variants encoding multiple isoforms have been observed for this gene. [provided by RefSeq, Dec 2011]

Specifications

Catalog Number	KC-3785
Cell Line Name	K562-41BBL-mem-IL21 Cell Line
NCBI/UniProt Accession Number	NM_003811.4 & NM_021803.4
Clone Number	NA
Host Cell Line	K562
Description	Stable K562 cell line expressing exogenous 41BBL and mem-IL21 gene
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+1µg/mL Puromycin
Selection Marker	Puromycin
Morphology	Lymphoblast
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
Doubling Time	Approximately 30-40 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

K562-41BBL-mem-IL21 cell line was generated using lentivirus expressing 41BBL-mem-IL21 sequence.

Characterization

Figure 1. Characterization of 41BBL over-expression in the K562-41BBL-mem-IL21 stable clone using FACS.

Figure 2. Characterization of Flag (mem-IL21) over-expression in the K562-41BBL-mem-IL21 stable clone using FACS.

Figure 3. In vitro expansion and phenotypic analysis of NK cells supported by K562-41BB6-mem-IL21 feeder cells.

Figure 4. Characterization of mem-IL21 and 41BBL over-expression in the K562-41BBL-mem-IL21 stable clone using PCR.

Cell Resuscitation

Prewarm culture medium (RPMI1640 supplemented with 10% FBS and 1μg/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 1:4-1:5 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

Prepare the freezing medium (70% RPMI1640 + 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

Zeng Q, Zhou Y, Schwarz H. CD137L-DCs, Potent Immune-Stimulators-History, Characteristics, and Perspectives. Front Immunol. 2019 Oct 2;10:2216. doi: 10.3389/fimmu.2019.02216. PMID: 31632390; PMCID: PMC6783506.
Zhao XY, Lv M, Xu LL, Qian X, Huang XJ. Donor Th17 cells and IL-21 may contribute to the development of chronic graft-versus-host disease after allogeneic transplantation. Eur J Immunol. 2013 Mar;43(3):838-50. doi: 10.1002/eji.201242816. Epub 2013 Jan 31. PMID: 23280509.

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.