KC-5075

293T-rat-ADAM9-Low Cell Line

55373

Home » 293T-rat-ADAM9-Low Cell Line

Background of 293T-rat-ADAM9-Low Cell Line

ADAM9 is a member of the ADAM family, a multifunctional and multi-domain type 1 transmembrane protein, and a membrane-anchored protein. It is expressed in various cell types, including monocytes and macrophages, and participates in multiple physiological functions mainly through its adhesive protease domain and metalloproteinase domain for shedding various cell surface proteins. ADAM9 is overexpressed in many cancers, including non-small cell lung cancer, pancreatic cancer, gastric cancer, breast cancer, ovarian cancer, and colorectal cancer, but is expressed limitedly in normal tissues. It is also associated with tumor invasiveness and poor prognosis.

Specifications

Catalog Number	KC-5075
Cell Line Name	293T-rat-ADAM9-Low Cell Line
Host Cell Line	293T
Description	Stable HEK293T clone expressing exogenous rat-ADAM9-Low 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% DMEM + 20% FBS + 10% DMSO
Propagation Medium	DMEM + 10% FBS + 1μg/mL Puromycin
Selection Marker	Puromycin
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-rat-ADAM9-Low cell line was generated using a lentiviral vector expressing the rat-ADAM9-Low sequence.

Characterization

Figure 1: Characterization of rat-ADAM9-Low in the 293T rat-ADAM9-Low stable clone using qPCR sequencing.

Figure 2: Characterization of rat-ADAM9-Low overexpression in the 293T rat-ADAM9-Low stable clone using PCR sequence

Cell Resuscitation

1. Prewarm culture medium (DMEM supplemented with 10% FBS and 1μg/mL puromycin)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. Chou CW, Huang YK, Kuo TT, Liu JP, Sher YP. An Overview of ADAM9: Structure, Activation, and Regulation in Human Diseases. Int J Mol Sci. 2020 Oct21;21(20):7790. doi: 10.3390/ijms21207790. PMID: 33096780; PMCID: PMC7590139.
2. Bazzone LE, Zhu J, King M, Liu G, Guo Z, MacKay CR, Kyawe PP, Qaisar N, Rojas-Quintero J, Owen CA, Brass AL, McDougall W, Baer CE, Cashman T, Trivedi CM, Gack MU, Finberg RW, Kurt-Jones EA. ADAM9 promotes type I interferon- mediated innate immunity during encephalomyocarditis virus infection. Nat Commun. 2024 May 16;15(1):4153. doi: 10.1038/s41467-024-48524-6. PMID: 38755212;PMCID: PMC11098812.
3.Umeda M, Yoshida N, Hisada R, Burbano C, Orite SYK, Kono M, Kyttaris VC, Krishfield S, Owen CA, Tsokos GC. ADAM9 enhances Th17 cell differentiation and autoimmunity by activating TGF-β1. Proc Natl Acad Sci U S A. 2021 May 4;118(18):e2023230118. doi: 10.1073/pnas.2023230118. PMID: 33911034; PMCID: PMC8106302.