KC-1606

293T-ACEII Cell Line

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Background of 293T-ACEII Cell Line

ACEII is also known as ACEH, is an enzyme that belongs to the angiotensin-converting enzyme family of dipeptidyl carboxydipeptidases and has considerable homology to human angiotensin 1 converting enzyme. ACE2 is known to be expressed in various human organs, and its organ- and cell-specific expression suggests that it may play a role in the regulation of cardiovascular and renal function, as well as fertility. In addition, ACEII is a functional receptor for the spike glycoprotein of the human coronavirus HCoV-NL63 and the human severe acute respiratory syndrome coronaviruses, SARS-CoV and SARS-CoV-2, the latter is the causative agent of coronavirus disease-2019(COVID-19).

Specifications

Catalog Number	KC-1606
Cell Line Name	293T-ACEII Cell Line
Host Cell Line	293T
Description	Stable 293T cell line expressing exogenous ACEII 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 + 1μg/mL Puromycin
Selection Marker	Puromycin
Morphology	Epithelial
Subculture	Split saturated culture 1:4-1:5 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
In Vivo Validation	NA

Cell Line Generation

293T-ACEII cell line was generated using a lentiviral vector expressing the ACEII sequence.

Characterization

Figure 1: Characterization of ACEII overexpression in the 293T-ACEII stable clone using qPCR.

Cell Resuscitation

Prewarm culture medium (DMEM supplemented with 10% FBS and 1μg/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 1:4-1:5 every 2-3 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

"Gene: ACE2, angiotensin I converting enzyme 2". National Center for Biotechnology Information (NCBI). U.S. National Library of Medicine. 2020-02-28.
Hikmet F, Méar L, Edvinsson Å, Micke P, Uhlén M, Lindskog C (July 2020). "The protein expression profile of ACE2 in human tissues". Molecular Systems Biology. 16 (7): e9610. doi:10.15252/msb.20209610. PMC 7383091. PMID 32715618.
Hamming I, Timens W, Bulthuis ML, Lely AT, Navis G, van Goor H (June 2004). "Tissue distribution of ACE2 protein, the functional receptor for SARS coronavirus. A first step in understanding SARS pathogenesis". The Journal of Pathology. 203 (2): 631–637. doi:10.1002/path.1570. PMC 7167720. PMID 15141377.
Kasmi Y, Khataby K, Souiri A (2019). "Coronaviridae: 100,000 Years of Emergence and Reemergence". In Ennaji MM (ed.). Emerging and Reemerging Viral Pathogens: Fundamental and Basic Virology Aspects of Human, Animal and Plant Pathogens. Vol. 1. Elsevier. p. 135. ISBN 978-0-12-819400-3.
Donoghue M, Hsieh F, Baronas E, Godbout K, Gosselin M, Stagliano N, et al. (September 2000). "A novel angiotensin-converting enzyme-related carboxypeptidase (ACE2) converts angiotensin I to angiotensin 1-9". Circulation Research. 87 (5): E1–E9.

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.