In Vitro Technologies

Permits and Restrictions	View Permits
Organism	Homo sapiens, human
Tissue	kidney
Morphology	epithelial
Culture Properties	adherent
Biosafety Level	2 [Cells contain Adeno and SV-40 viral DNA sequences] Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country.
Age	embryo
Applications	Phoenix is a second-generation retrovirus producer cell line for the generation of helper-free ecotropic and amphotropic retroviruses. The unique feature of this cell line is that it is highly transfectable with either calcium phosphate mediated transfection or lipid-based transfection protocols-- up to 50% or higher of cells can be transiently transfected. This cell line is capable of carrying episomes for long-term stable production of retrovirus (LZRS).
Storage Conditions	liquid nitrogen vapor phase

Derivation	HEK 293T/17 cells were transformed with adenovirus E1a carrying a temperature sensitive T antigen co-selected with neomycin. Transformation was brought about by the insertion of approximately 4.5 kilobases of viral genome into human chromosome 19. Gag-pol was introduced with hygromycin as the co-selectable marker and the envelope proteins were introduced with diptheria resistance as the co-selectable marker.
Comments	Phoenix Ecotropic (ECO) cell line expresses ecotropic envelope protein. CD8 expression is a direct reflection of intracellular gag-pol and the stability of the producer cell population's ability to produce gag-pol can be readily monitored by flow cytometry. Phoenix-ECO have been extensively tested for helper virus production and established as being helper-virus free.

Derivation

HEK 293T/17 cells were transformed with adenovirus E1a carrying a temperature sensitive T antigen co-selected with neomycin. Transformation was brought about by the insertion of approximately 4.5 kilobases of viral genome into human chromosome 19. Gag-pol was introduced with hygromycin as the co-selectable marker and the envelope proteins were introduced with diptheria resistance as the co-selectable marker.

Comments

Phoenix Ecotropic (ECO) cell line expresses ecotropic envelope protein.

CD8 expression is a direct reflection of intracellular gag-pol and the stability of the producer cell population's ability to produce gag-pol can be readily monitored by flow cytometry.

Phoenix-ECO have been extensively tested for helper virus production and established as being helper-virus free.

Complete Growth Medium	The base medium for this cell line is Dulbecco’s Modified Eagle’s Medium (DMEM) (ATCC 30-2002). To make the complete growth medium, add the following components to the base medium: 10% Fetal Bovine Serum (heat inactivated) (ATCC 30-2020), 2mM L-glutamine (ATCC 30-2214), 1% Penicillin/Streptomycin.
Subculturing	Remove and discard culture medium. Briefly rinse the cell layer with 0.25% (w/v) Trypsin- 0.53 mM EDTA solution to remove all traces of serum that contains trypsin inhibitor. Add 2.0 to 3.0 mL of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until cell layer is dispersed (usually within 5 to 15 minutes). Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Cells that are difficult to detach may be placed at 37°C to facilitate dispersal. Add 6.0 to 8.0 mL of complete growth medium and aspirate cells by gently pipetting. Add appropriate aliquots of the cell suspension to new culture vessels. Incubate cultures at 37°C. Note: Culture to ≤80% confluence to prevent the cells from sloughing Subcultivation Ratio: A subcultivation ratio of 1:5 to 1:8 is recommended Medium Renewal: Every 2 to 3 days
Cryopreservation	Freeze medium: complete growth medium supplemented with 5% (v/v) DMSO Storage temperature: liquid nitrogen vapor phase
Culture Conditions	Temperature: 37°CAtmosphere: air, 95%; carbon dioxide (CO₂), 5%

STR Profile	D5S818: 8, 9 D13S317: 11, 12, 13 D7S820: 11 D16S539: 9, 13 vWA: 16, 19 TH01: 7, 9.3 Amelogenin: X TPOX: 11 CSF1PO: 11, 12
Population Doubling Time	16 hours

Name of Depositor	G Nolan
Year of Origin	1996
References	Swift S et al. Rapid Production of Retroviruses for Efficient Gene Delivery to Mammalian Cells Using 293T Cell-Based Systems. Current Protocols in Immunology,Unit 10.17C, 2001 Pear WS, et al. Production of High-Titer Helper-Free Retroviruses by Transient Transfection. Proc. Natl. Acad. Sci. USA 90: 8392-8396, 1993. PubMed: 7690960 Nolan Laboratory Retroviral Systems. Phoenix helper-free retrovirus producer lines, https://web.stanford.edu/group/nolan/_OldWebsite/retroviral_systems/retsys.html

Name of Depositor

G Nolan

Year of Origin

1996

References

Swift S et al. Rapid Production of Retroviruses for Efficient Gene Delivery to Mammalian Cells Using 293T Cell-Based Systems. Current Protocols in Immunology,Unit 10.17C, 2001

Pear WS, et al. Production of High-Titer Helper-Free Retroviruses by Transient Transfection. Proc. Natl. Acad. Sci. USA 90: 8392-8396, 1993. PubMed: 7690960

Nolan Laboratory Retroviral Systems. Phoenix helper-free retrovirus producer lines, https://web.stanford.edu/group/nolan/_OldWebsite/retroviral_systems/retsys.html

Permits	These permits may be required for shipping this product to Australia: DAFF Import Permit formerly known as AQIS Import Permit must be obtained and a copy of the permit must be sent to ATCC in advance of shipment.
Basic Documentation	Product Sheet Certificate of Analysis SDS
References	Swift S et al. Rapid Production of Retroviruses for Efficient Gene Delivery to Mammalian Cells Using 293T Cell-Based Systems. Current Protocols in Immunology,Unit 10.17C, 2001 Pear WS, et al. Production of High-Titer Helper-Free Retroviruses by Transient Transfection. Proc. Natl. Acad. Sci. USA 90: 8392-8396, 1993. PubMed: 7690960 Nolan Laboratory Retroviral Systems. Phoenix helper-free retrovirus producer lines, https://web.stanford.edu/group/nolan/_OldWebsite/retroviral_systems/retsys.html

Permits

These permits may be required for shipping this product to Australia:

DAFF Import Permit formerly known as AQIS Import Permit must be obtained and a copy of the permit must be sent to ATCC in advance of shipment.

Basic Documentation

Product Sheet Certificate of Analysis SDS

References

Swift S et al. Rapid Production of Retroviruses for Efficient Gene Delivery to Mammalian Cells Using 293T Cell-Based Systems. Current Protocols in Immunology,Unit 10.17C, 2001

Pear WS, et al. Production of High-Titer Helper-Free Retroviruses by Transient Transfection. Proc. Natl. Acad. Sci. USA 90: 8392-8396, 1993. PubMed: 7690960

Nolan Laboratory Retroviral Systems. Phoenix helper-free retrovirus producer lines, https://web.stanford.edu/group/nolan/_OldWebsite/retroviral_systems/retsys.html

E: care@invitro.com.au
P: 1300 552 003

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Phoenix-ECO; Embryonic Kidney Cells; Human (Homo sapiens) Item ID: ATCCRL3214

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