KC-5693

293T-MECA447-P Cell Line

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Background of 293T-MECA447-P Cell Line

CD19 (CD19 Molecule) is a Protein Coding gene. This gene encodes a member of the immunoglobulin gene superfamily. CD19 is a B-cell transmembrane molecule that is critical for B-cell activation. Expression of this cell surface protein is restricted to B cell lymphocytes. This protein is a reliable marker for pre-B cells but its expression diminishes during terminal B cell differentiation in antibody secreting plasma cells. Activation of B lymphocytes involves binding of antigen to the specific receptor and signalling through several membrane coreceptors, of which CD19 has been found to play a pivotal role as a response regulator. MECA447 is an autoimmune-associated antigen, MECA447 may be related to autoantigens associated with vascular endothelial cells and is often used in rheumatology and immunology research. Diseases associated with CD19 include Immunodeficiency, Common Variable, 3 and Common Variable Immunodeficiency Phenotype Due To Cd19/Cd81 Deficiency.

Specifications

Catalog Number	KC-5693
Cell Line Name	293T-MECA447-P Cell Line
Clone Number	3#
Host Cell Line	293T
Description	Stable 293T clone expressing exogenous human MECA447-P gene
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% DMEM + 20% FBS + 10% DMSO
Propagation Medium	DMEM + 10% FBS + 150μg/mL Hygromycin B
Selection Marker	Hygromycin B
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 30 hours
Mycoplasma Status	Negative

Cell Line Generation

293T-human-MECA447-P cell line was generated using a lentiviral vector expressing the human MECA447-P sequence.

Characterization

Figure 1: Characterization of mouse NRP1 overexpression in the 293T human MECA447-P stable clone using FACS(Kyinno, Cat#KP-1123).

Figure 2: Characterization of human MECA447-P in the 293T stable clone using PCR sequencing.

Cell Resuscitation

1. Prewarm culture medium (DMEM + 10% FBS + 150μg/mL Hygromycin B)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% DMEM + 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. Fujimoto M, Poe JC, Jansen PJ, Sato S, Tedder TF. CD19 amplifies B lymphocyte signal transduction by regulating Src-family protein tyrosine kinase activation. J Immunol. 1999 Jun 15;162(12):7088-94. PMID: 10358152.
2. Gärdby E, Chen XJ, Lycke NY. Impaired CD40-signalling in CD19-deficient mice selectively affects Th2-dependent isotype switching. Scand J Immunol. 2001 Jan;53(1):13-23. doi: 10.1046/j.1365-3083.2001.00824.x. PMID: 11169202.
3. Kong D, Kwon D, Moon B, Kim DH, Kim MJ, Choi J, Kang KS. CD19 CAR-expressing iPSC-derived NK cells effectively enhance migration and cytotoxicity into glioblastoma by targeting to the pericytes in tumor microenvironment. Biomed Pharmacother. 2024 May;174:116436. doi: 10.1016/j.biopha.2024.116436. Epub 2024 Mar 19. PMID: 38508081.

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.