KC-6062

Jurkat-NFAT-Luc2-LY6E-KO-CD16a-V158 Cell Line

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Background of Jurkat-NFAT-Luc2-LY6E-KO-CD16a-V158 Cell Line

The lymphocyte antigen 6 family member E (LY6E) gene is localized to human chromosome 8q24.3 (143,017,982–143,023,832 bp) and encodes a glycosylphosphatidylinositol (GPI)-anchored cell surface protein. As a member of the LY6 superfamily, it is widely expressed in the liver, kidney, ovary, spleen, and peripheral blood leukocytes, with critical roles in immune regulation, T-cell development, and signal transduction. Aberrant overexpression of LY6E is observed in multiple malignancies including breast, lung, gastric, and ovarian cancers, and is closely associated with poor patient survival and tumor immune escape. Its restricted expression in normal tissues and oncogenic functions make it a promising target for antibody-drug conjugates (ADCs) and other anticancer therapies. Additionally, LY6E acts as an interferon-stimulated gene, participating in host responses to viral infections.

Specifications

Catalog Number	KC-6062
Cell Line Name	Jurkat-NFAT-Luc2-LY6E-KO-CD16a-V158 Cell Line
NCBI/UniProt Accession Number	4061
Clone Number	2A3
Host Cell Line	Jurkat-NFAT-Luc2-CD16a-V158
Description	Stable Jurkat-NFAT-Luc2-CD16a-V158 cell line with LY6E gene knockout, No.2A3
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+300μg/mL Hygromycin B+0.75μg/mL Puromycin
Selection Marker	Puromycin \| Hygromycin B
Morphology	Lymphoblast
Subculture	Split saturated culture 1:4-1:6 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 26 hours
Mycoplasma Status	Negative

Cell Line Generation

Jurkat-NFAT-Luc2-LY6E-KO-CD16a-V158 cell line was generated using the CRISPR method.

Characterization

Figure 1: Characterization of Jurkat-NFAT-Luc2-LY6E-KO-CD16a-V158 Cell Line stable clone using PCR sequencing.

Figure 2: Characterization of Jurkat-NFAT-Luc2-LY6E-KO-CD16a-V158 Cell Line stable clone using RT-PCR sequencing.

Figure 3: Characterization of Jurkat-NFAT-Luc2-LY6E-KO-CD16a-V158 Cell Line stable clone using Western blot.

Cell Resuscitation

Prewarm culture medium (RPMI1640+10% FBS+300μg/mL Hygromycin B+0.75μg/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 1-3 × 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

Luo L, McGarvey P, Madhavan S, Kumar R, Gusev Y, Upadhyay G. Distinct lymphocyte antigens 6 (Ly6) family members Ly6D, Ly6E, Ly6K and Ly6H drive tumorigenesis and clinical outcome. Oncotarget. 2016 Mar 8;7(10):11165-93. doi: 10.18632/oncotarget.7163. PMID: 26862846; PMCID: PMC4905465.
Mao M, Yu M, Tong JH, Ye J, Zhu J, Huang QH, Fu G, Yu L, Zhao SY, Waxman S, Lanotte M, Wang ZY, Tan JZ, Chan SJ, Chen Z. RIG-E, a human homolog of the murine Ly-6 family, is induced by retinoic acid during the differentiation of acute promyelocytic leukemia cell. Proc Natl Acad Sci U S A. 1996 Jun 11;93(12):5910-4. doi: 10.1073/pnas.93.12.5910. PMID: 8650192; PMCID: PMC39161.