KC-5639

CHOK1-rat-IL31RA-Flag-Low Cell Line

59954

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Background of CHOK1-rat-IL31RA-Flag-Low Cell Line

IL31RA (Interleukin 31 Receptor A) is a Protein Coding gene. The protein encoded by this gene belongs to the type I cytokine receptor family. This receptor, with homology to gp130, is expressed on monocytes, and is involved in IL-31 signaling via activation of STAT-3 and STAT-5. Type 2 inflammatory cytokines such as interleukin (IL)-4, IL-13, and IL-31 are strongly implicated in AD pathogenesis. Stimulation of IL-31 cognate receptors on C-fiber nerve endings is believed to activate neurons in the dorsal root ganglion (DRG), causing itch. The IL-31 receptor is a heterodimer of OSMRβ and IL31RA subunits, and OSMRβ can also bind oncostatin M (OSM), a pro-inflammatory cytokine released by monocytes/macrophages, dendritic cells, and T lymphocytes. IL-31 receptor α-chain (IL-31RA) is also expressed in peripheral nerves and epidermal keratinocytes, and the roles of IL-31 on pruritus and skin barrier have been investigated.

Specifications

Catalog Number	KC-5639
Cell Line Name	CHOK1-rat-IL31RA-Flag-Low Cell Line
NCBI/UniProt Accession Number	NM_001257278.2
Clone Number	23#
Host Cell Line	CHOK1
Description	Stable CHOK1 clone expressing exogenous rat IL31RA Flag gene in low level
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 + 10μg/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 24 hours
Mycoplasma Status	Negative

Cell Line Generation

CHOK1-rat-IL31RA-Flag-Low cell line was generated using a lentiviral vector expressing the rat IL31RA Flag sequence.

Characterization

Figure 1: Characterization of endogenous Flag tag expression in CHOK1 cell using FACS.

Figure 2: Characterization of Flag tag overexpression in the CHOK1-rat-IL31RA-Flag-Low stable clone using FACS.

Figure 3: Characterization of rat IL31RA in the CHOK1-rat-IL31RA-Flag-Low stable clone using PCR sequencing.

Cell Resuscitation

1. Prewarm culture medium (RPMI1640 supplemented with 10% FBS and 10μg/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% 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. Datsi A, Steinhoff M, Ahmad F, Alam M, Buddenkotte J. Interleukin-31: The "itchy" cytokine in inflammation and therapy. Allergy. 2021 Oct;76(10):2982-2997. doi: 10.1111/all.14791. Epub 2021 Mar 16. PMID: 33629401.
2. Suehiro M, Numata T, Saito R, Yanagida N, Ishikawa C, Uchida K, Kawaguchi T, Yanase Y, Ishiuji Y, McGrath J, Tanaka A. Oncostatin M suppresses IL31RA expression in dorsal root ganglia and interleukin-31-induced itching. Front Immunol. 2023 Nov 16;14:1251031. doi: 10.3389/fimmu.2023.1251031. PMID: 38035099; PMCID: PMC10687395.
3. Nakashima C, Otsuka A, Kabashima K. Interleukin-31 and interleukin-31 receptor: New therapeutic targets for atopic dermatitis. Exp Dermatol. 2018 Apr;27(4):327-331. doi: 10.1111/exd.13533. PMID: 29524262.

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.