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Cell Biology
ATCC® Number: CRL-2782™    Price: $338.00
Designations: 293 EcR Shh [JHU-64]
Depositors:  PA Beachy
Biosafety Level: 2 [Cells containing Adenovirus V viral sequences ]
Shipped: frozen
Medium & Serum: See Propagation
Growth Properties: adherent
Organism: Homo sapiens (human)
Morphology: epithelial
Source: Organ: kidney
Cell Type: epithelialtransformed with adenovirus 5 DNA
Permits/Forms: In addition to the MTA mentioned above, other ATCC and/or regulatory permits may be required for the transfer of this ATCC material. Anyone purchasing ATCC material is ultimately responsible for obtaining the permits. Please click here for information regarding the specific requirements for shipment to your location.
Restrictions: Part of the Johns Hopkins Special Collection
Isolation: Isolation date: 1998
DNA Profile (STR): Amelogenin: X
CSF1PO: 12
D13S317: 12,14
D16S539: 9,13
D5S818: 8
D7S820: 11,12
THO1: 7,9.3
TPOX: 11
vWA: 16,19
Age: fetus
Comments: 293 EcR Shh cells were created in 1998 by stably transfecting HK293 cells with the Ecdysone-Inducible Mammalian Expression System (Invitrogen). 293 EcR Shh cells carry a stably integrated construct for the expression of murine Sonic hedgehog (Shh) under ecdysone-inducible control. The Shh expressed in these cells is efficiently processed, undergoing both internal cleavage and the covalent addition of both cholesterol (carboxy-terminus) and a fatty acid (amino-terminus) to generate the mature signaling molecule. 293 EcR Shh cells can therefore be used for the production of biologically active murine Shh.
Propagation: ATCC complete growth medium: Dulbecco's modified Eagle's medium with 4 mM L-glutamine adjusted to contain 1.5 g/L sodium bicarbonate and 4.5 g/L glucose supplemented with 0.4 mg/ml G-418, 0.4 mg/ml Zeocin (Invitrogen, Cat. No. R25001) and 10% fetal bovine serum
Atmosphere: air, 95%; carbon dioxide (CO2), 5%
Temperature: 37.0°C
Subculturing: Protocol:
  1. Remove and discard culture medium.
  2. 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.
  3. 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.
  4. Add 6.0 to 8.0 ml of complete growth medium and aspirate cells by gently pipetting.
  5. Add appropriate aliquots of the cell suspension to new culture vessels.
    An inoculum of 8 X 10(3) to 8 X 10(4) viable cells/cm2 is recommended. Maintain cultures at a cell concentration between 2 X 10(4) and 2 X 10(5) cells/cm2. Do not exceed 5 X 10(5) cells/cm2.
  6. Incubate cultures at 37°C.

Subcultivation Ratio: A subcultivation ratio of 1:4 to 1:10 is recommended
Medium Renewal: Two to three times weekly
Preservation: Freeze medium: Complete growth medium supplemented with 5% (v/v) DMSO
Storage temperature: liquid nitrogen vapor phase
Doubling Time: 23 hrs
Related Products: Recommended medium (without the additional supplements or serum described under ATCC Medium):ATCC 30-2002
recommended serum:ATCC 30-2020
source culture:ATCC JHU-64
References: 71396: Taipale J, et al. Effects of oncogenic mutations in Smoothened and Patched can be reversed by cyclopamine. Nature 406: 1005-1009, 2000. PubMed: 10984056
71398: Cooper MK, et al. Teratogen-mediated inhibition of target tissue response to Shh signaling. Science 280: 1603-1607, 1998. PubMed: 9616123
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