KC-1676

293T-OS8-PVR-Cell-Line

22595

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Background of 293T-OS8-PVR-Cell-Line

PVR, abbreviated as poliovirus receptor, is a transmembrane glycoprotein belonging to immunoglobulin superfamily. CD155 is involved in the cellular poliovirus infection in primates, and intercellular adherent junctions between epithelial cells.

Specifications

Catalog Number	KC-1676
Cell Line Name	293T-OS8-PVR-Cell-Line
Host Cell Line	Human HEK293T cell line
Description	Stable HEK293T cell line expressing exogenous human PVR gene and OS8 sequence (a ScFV sequence of anti-human CD3 mAb OKT3 and a C-terminal domain of mouse CD8a which consist of transmembrane and cytoplasmic domains).
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% DMEM+20% FBS+10% DMSO
Propagation Medium	DMEM+10%FBS+1µg/mL Puromycin+150µg/mL Hygromycin B
Selection Marker	Puromycin, HygromycinB
Morphology	Epithelial
Subculture	Split saturated culture 1:3-1:6 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

293T OS8 human PVR cell line was generated using a lentiviral vector expressing the human PVR and OS8 sequence (a ScFV sequence of anti-human CD3 mAb OKT3 and a C-terminal domain of mouse CD8a which consist of transmembrane and cytoplasmic domains)..

Characterization

Cell Resuscitation

Prewarm culture medium (DMEM + 10% FBS + 1µg/mL Puromycin + 150µg/mL Hygromycin B)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 1:3-1:6 every 3 days; seed out at about 1-3 × 10⁵ cells/mL.

Cell Freezing

Prepare the freezing medium (70% DMEM + 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

Koike S, Horie H, Ise I, Okitsu A, Yoshida M, Iizuka N, Takeuchi K, Takegami T, Nomoto A (October 1990). "The poliovirus receptor protein is produced both as membrane-bound and secreted forms". EMBO J. 9 (10): 3217ÿ 24.
Mendelsohn CL, Wimmer E, Racaniello VR (1989). "Cellular receptor for poliovirus: molecular cloning, nucleotide sequence, and expression of a new member of the immunoglobulin superfamily". Cell. 56 (5): 855ÿ 65.
Maier MK, Seth S, Czeloth N, et al. (2007). "The adhesion receptor CD155 determines the magnitude of humoral immune responses against orally ingested antigens". European Journal of Immunology. 37 (8): 2214ÿ25.