These cells constitutively express the simian virus 40 (SV40) large T antigen, and clone 17 was selected specifically for its high transfectability.  CRL-11268 293T/17人胚腎細(xì)胞

SV40 T antigen

SV40 T antigen

293T/17 cells were cotransfected with the pCRIPenv- and the pCRIPgag-2 vectors to obtain the ANJOU 65 (see ATCC CRL-11269) cell line. ANJOU 65 cells were cotransfected with the pCRIPgag-2 and pGPT2E vectors to obtain the BOSC 23 (see ATCC CRL-11270) ecotropic envelope-expression packaging cell line. ANJOU 65 cells were also cotransfected with the pCRIPAMgag vector along with a plasmid expressing the gpt resistance gene to obtain the Bing (see ATCC CRL-11554)amphotropic envelope-expression packaging cell line.

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.

6. Incubate cultures at 37°C.

S

Medium Renewal: Every 2 to 3 days

Freeze medium: Complete growth medium supplemented with 5% (v/v) DMSO

Storage temperature: liquid nitrogen vapor phase

Temperature: 37°C

Atmosphere: air, 95%; carbon dioxide (CO2), 5%