NIH3T3 Cell Line

NIH3T3 Cell Line General Information Organism Mus musculus, mouse Cell Line NIH 3T3 mouse embryonic fibroblast cells were initiated from a cell Description line isolated at the New York University School of Medicine Department of Pathology in 1962. 3T3 refers to the cell transfer and inoculation method for the line, and means “3-day transfer, inoculum 3 x 105 cells.” Using this method, the immortal cell line begins to thrive and stabilize in cell culture after about 20-30 generations of in vitro growth. George Todaro and Howard Green originally obtained the cells from desegregated NIH Swiss mouse embryo fibroblasts. Strain NIH/Swiss Tissue Embryo Morphology Fibroblast Age Embryo Product Frozen Format Growth Mode Adherent

Biosafety 1 Level 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. Applications DNA transfection studies NIH 3T3 cell line has been used to: 1. study calcium-mediated actin reset (CaAR) in response to physiological changes 2. determine the viral titer of the retroviral construct - ST40hTNFα 3. study the effects of activation of the transient receptor potential vanilloid 3 channel (TRPV3) on adipocyte differentiation Shipped in Dry ice −196°C Storage Temperature Characteristics Karyotype Not specified Images Products Not specified

Receptors Not specified Mycoplasma Contamination was eliminated with Mycoplasma Removal Agent, then negative in DAPI, microbiological culture, PCR assays, RNA hybridization Cytogenetics Murine hypertriploid karyotype with 3% polyploidy - 68(58-73) <3n> Viruses ELISA: reverse transcriptase negative; PCR: SMRV - Comments The original cells are extremely contact inhibited, although the cell line is no longer inhibited. NIH 3T3 cells are receptive to transformation with SV40 and polyomavirus. Moreover, 3T3 cells are sensitive to sarcoma virus focus formation, leukemia virus and are inhibited by temazepam and other benzodiazepines. Culture Conditions and Handling Culture DMEM + 2 mM Glutamine + 10% Calf Serum (CS). Medium Subculturing 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 2.0-3.0 mL of Trypsin-EDTA solution to flask and observe cells under an inverted microscope until the cell layer is dispersed. 4. Add 6.0-8.0 mL of complete growth medium, and then aspirate cells by gently pipetting. 5. Add appropriate aliquots of the cell suspension to the new culture vessel. 6. Incubate cultures at 37°C. Split Ratio Split culture at 1:10 every 4 to 6 days using trypsin/EDTA

Medium Twice per week Renewal Culture 37°C Temperature Incubation Carbon dioxide (CO2), 5% Condition Doubling Time Ca. 20 hours Harvest Cell harvest of about 20 x 106 cells/175 cm2 flask Storage Frozen with 70% medium, 20% FBS, 10% DMSO