KC-1500

THP-1-siglec15 Cell Line

22261

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Background of THP-1-siglec15 Cell Line

Siglec-15 is a transmembrane glycoprotein, which belongs to the immunoglobulin superfamily and SIGLEC (sialic acid binding Ig-like lectin) family, and is found primarily on the surface of immune cells. Siglec-15 is that recognizes sialylated glycans and regulates osteoclast differentiation. Siglec-15 is a potential therapeutic target for osteoporosis and plays a conserved regulatory role in the immune system of vertebrates.

Specifications

Catalog Number	KC-1500
Cell Line Name	THP-1-siglec15 Cell Line
Host Cell Line	THP-1
Description	THP-1 cell line stable expressing exogenous human siglec15 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+2µg/mL Puromycin
Selection Marker	Puromycin
Morphology	Monocyte
Subculture	Split saturated culture 1:4-1:8 every 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 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

THP-1 human siglec15 cell line was generated using lentiviral vector expressing human siglec15 sequence.

Characterization

Figure 1: Characterization of siglec15 overexpression in the THP-1 siglec15 stable clone using FACS.

Cell Resuscitation

Prewarm culture medium (RPMI1640 + 10% FBS + 2µg/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 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

Wang, Jun, Jingwei Sun, Linda N Liu, Dallas B Flies, Xinxin Nie, Maria Toki, Jianping Zhang, et al. 2019. ¡ùSiglec15 as an Immune Suppressor and Potential Target for Normalization Cancer Immunotherapy.¡ì Nature Medicine, March. Springer US, 1Ã¿32. doi:10.1038/s41591-019-0374-x.