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HEK 293 STF  拉丁名

ATCC CRL-3249 編號

Product category Human cells 

Product type Reporter-labeled cell 

Organism Homo sapiens, human 

Cell type epithelial-like cell 

Morphology epithelial-like 

Tissue kidney; Embryo 

Applications 3D cell culture Assay development High-throughput screening 

Product format 提供形式 Frozen 

Storage conditions 保藏條件 Vapor phase of liquid nitrogen

Specific applications Assaying canonical Wnt signaling pathway. 

This is a luciferase reporter cell line that responds to canonical Wnt signaling by expressing firefly luciferase.Growth properties Adherent Age fetus 

Comments This cell line was stably cotransfected with (1) a 7 x LEF/TCF sites, a minimal thymidine kinase promoter and (2) a pSV2-neo resistance plasmid exhibiting G418 resistance. 

Luciferase activity reflects the level of canonical Wnt signaling.

Unpacking and storage instructions Check all containers for leakage or breakage. 

Remove the frozen cells from the dry ice packaging and immediately place the cells at a temperature below --130°C, preferably in liquid nitrogen vapor, until ready for use. 

Complete medium The base medium for this cell line is DMEM F12 Medium, (ATCC? 30-2006?), 80%. 

To make the complete growth medium, add the following components to the base medium: 200 μg/mL G-418 Bovine Calf Serum, Iron Fortified (ATCC? 30-2030?), 20% 

Temperature 培養(yǎng)溫度 37°C 

Atmosphere 需氧情況 95% Air, 5% CO2 

Handling procedure To insure the highest level of viability, thaw the vial and initiate the culture as soon as possible upon receipt. If upon arrival, continued storage of the frozen culture is necessary, it should be stored in liquid nitrogen vapor phase and not at -70°C. Storage at -70°C will result in loss of viability. 

Thaw the vial by gentle agitation in a 37°C water bath. 

To reduce the possibility of contamination, keep the O-ring and cap out of the water.  

Thawing should be rapid (approximately 2 minutes). Remove the vial from the water bath as soon as the contents are thawed, and decontaminate by dipping in or spraying with 70% ethanol. 

All of the operations from this point on should be carried out under strict aseptic conditions. 

Transfer the vial contents to a centrifuge tube containing 9.0 mL complete growth medium and spin at approximately 125 x g for 5 to7 minutes. 

Resuspend cell pellet with the recommended complete growth medium (see the specific batch information for the culture recommended dilution ratio) and dispense into a 25 cm2 or a 75 cm2 culture flask. 

It is important to avoid excessive alkalinity of the medium during recovery of the cells. 

It is suggested that, prior to the addition of the vial contents, the culture vessel containing the complete growth medium be placed into the incubator for at least 15 minutes to allow the medium to reach its normal pH (7.0 to 7.6). 

Incubate the culture at 37°C in a suitable incubator. 

A 5% CO2 in air atmosphere is recommended if using the medium described on this product sheet. 

Subculturing procedure Volumes used in this protocol are for 75 cm2 flasks; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes. 

Remove and discard culture medium. Briefly rinse the cell layer with Ca++/Mg++ free Dulbecco's phosphate-buffered saline (D-PBS). Add 1.0 to 2.0 mL of 0.25% 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 37.0°C to facilitate dispersal. Add 6.0 to 8.0 mL of complete growth medium and aspirate cells by gently pipetting. 

Transfer cell suspension to a centrifuge tube and spin at approximately 125 x g for 5 to 10 minutes. 

Discard supernatant. Resuspend the cell pellet in fresh growth medium. Add appropriate aliquots of the cell suspension to new culture vessels. 

Subcultivation Ratio: A subcultivation ratio of 1:3 to 1:5 is recommended Medium Renewal: 2 to 3 times per week Seeding Density: 3.0 x 104 to 5.0x104 cells/cm2 

Reagents for cryopreservation Complete growth medium supplemented with 50% (v/v) calf serum and 10% (v/v) DMSO (ATCC 4-X)