KC-1774

CHOK1-ILDR2-Cell-Line

22775

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Background of CHOK1-ILDR2-Cell-Line

ILDR2 (Immunoglobulin Like Domain Containing Receptor 2) also known as C1orf32, it is a Protein Coding gene. ILDR2 is a novel B7-like protein that negatively regulates T cell responses. ILDR2 is a regulatory factor of diabetes susceptibility in obese mice. In vivo pharmacodynamic experiments, ILDR2 was expressed in most organs of islet and hypothalamus. Studies have shown that the manipulation of ILDR2 expression in the liver affects lipid homeostasis and ER stress pathways. Diseases associated with ILDR2 include Hypogonadotropic Hypogonadism 4 With Or Without Anosmia and Hypogonadotropic Hypogonadism 11 With Or Without Anosmia.

Specifications

Catalog Number	KC-1774
Cell Line Name	CHOK1-ILDR2-Cell-Line
Host Cell Line	CHOK1
Description	Stable CHOK1 cell line expressing exogenous ILDR2 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% 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
In Vivo Validation	NA

Cell Line Generation

CHOK1-ILDR2 Cell Line was generated using a lentiviral vector expressing the ILDR2 sequence.

Characterization

Figure 1: Characterization of ILDR2 overexpression in the CHO-K1 ILDR2 stable clone using FACS.

Cell Resuscitation

Prewarm culture medium (RPMI1640 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 1:4-1:8 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

Hecht I, Toporik A, Podojil JR, Vaknin I, Cojocaru G, Oren A, Aizman E, Liang SC, Leung L, Dicken Y, Novik A, Marbach-Bar N, Elmesmari A, Tange C, Gilmour A, McIntyre D, Kurowska-Stolarska M, McNamee K, Leitner J, Greenwald S, Dassa L, Levine Z, Steinberger P, Williams RO, Miller SD, McInnes IB, Neria E, Rotman G. ILDR2 Is a Novel B7-like Protein That Negatively Regulates T Cell Responses. J Immunol. 2018 Mar 15;200(6):2025-2037. doi: 10.4049/jimmunol.1700325. Epub 2018 Feb 5. PMID: 29431694; PMCID: PMC6860365.
Watanabe K, Watson E, Cremona ML, Millings EJ, Lefkowitch JH, Fischer SG, LeDuc CA, Leibel RL. ILDR2: an endoplasmic reticulum resident molecule mediating hepatic lipid homeostasis. PLoS One. 2013 Jun 24;8(6):e67234. doi: 10.1371/journal.pone.0067234. PMID: 23826244; PMCID: PMC3691114.