KC-1589

3A9-LILRB2-Cell-Line

22425

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Background of 3A9-LILRB2-Cell-Line

Leukocyte immunoglobulin-like receptor subfamily B member 2 (LILRB2), also known as ILT4, LIR2, monocyte/macrophage immunoglobulin-like receptor 10, MIR-10, and CD85d, belongs to the leukocyte immunoglobulin-like receptor (LIR) family of transmembrane glycoproteins that negatively regulate immune cell activation.LILRB2 is expressed on both innate (monocytes, macrophages, basophils, and DCs) and adaptive (CD4+ T cells) immune cells and interacts with MHC-I on nucleated cells (unknown specific ligand). LILRB2 interacts with related ligands HLA-G, ANGPTLs, SEMA4A and CD1d in tumor microenvironment, resulting in myeloid cells promoting tumor growth and enhancing tumor immune escape. It has been shown pre-clinically that LILRB2 antagonism promotes macrophage maturation and activation. MK-4830 (Merck and Agenus) is an anti-LILRB2 fully human IgG4 monoclonal antibody, and phase II clinical studies have been initiated. In addition, IO-108, JTX-8064, and NGM707 are in phase I clinical studies.

Specifications

Catalog Number	KC-1589
Cell Line Name	3A9-LILRB2-Cell-Line
NCBI/UniProt Accession Number	NM_005874.5
Host Cell Line	Mouse 3A9 cell line
Description	Stable 3A9 T cell hybridoma clone expressing exogenous LILRB2.
Quantity	One vial of frozen cells (≥2×10⁶ cells)
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-like
Subculture	Split saturated culture 1:4-1:6 every 2-3 days; seed out at about 1-2 × 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

3A9 human LILRB2 cell Line was generated using retrovirus vector expressing human LILRB2 sequence.

Characterization

Cell Resuscitation

Prewarm culture medium (RPMI1640 + 10% FBS + 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 1:4-1:6 every 2-3 days; seed out at about 1-2 × 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

Lentz RW, Colton MD, Mitra SS, Messersmith WA. Innate Immune Checkpoint Inhibitors: The Next Breakthrough in Medical Oncology? Mol Cancer Ther. 2021 Jun;20(6):961-974. doi: 10.1158/1535-7163.MCT-21-0041.
Chen HM, van der Touw W, Wang YS, Kang K, Mai S, Zhang J, Alsina-Beauchamp D, Duty JA, Mungamuri SK, Zhang B, Moran T, Flavell R, Aaronson S, Hu HM, Arase H, Ramanathan S, Flores R, Pan PY, Chen SH. Blocking immunoinhibitory receptor LILRB2 reprograms tumor-associated myeloid cells and promotes antitumor immunity. J Clin Invest. 2018 Dec 3;128(12):5647-5662. doi: 10.1172/JCI97570.
Zhao P, Xu Y, Jiang LL, Fan X, Ku Z, Li L, Liu X, Deng M, Arase H, Zhu JJ, Huang TY, Zhao Y, Zhang C, Xu H, Tong Q, Zhang N, An Z. LILRB2-mediated TREM2 signaling inhibition suppresses microglia functions. Mol Neurodegener. 2022 Jun 18;17(1):44. doi: 10.1186/s13024-022-00550-y.

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.