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Mesothelial cells were isolated from pleural fluids obtained from non-cancerous individuals.
The cells were transfected with the pRSV-T plasmid (an SV40 ori- construct containing the SV40 early region and the Rous sarcoma virus long terminal repeat) and cloned.
The cells can be used to screen chemical and biological agents for activity as growth factor, carcinogens, mutagens, etc.
The cells stain positively for vimentin, keratins and SV40 T antigen.
Propagation:
ATCC complete growth medium: Medium199 with Earle's BSS, .75 mM L-glutamine and 1.25 g/L sodium bicarbonate supplemented with with 3.3 nM epidermal growth factor (EGF), 400 nM hydrocortisone, 870 nM insulin, 20 mM HEPES, trace elements (see above reference) and 10% fetal bovine serum. Temperature: 37.0°C
Subculturing:
Protocol:
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.
To remove trypsin-EDTA solution, transfer cell suspension to centrifuge tube and spin at approximately 125 xg for 5 to 10 minutes.Discard supernatant and resuspend cells in fresh growth medium. Add appropriate aliquots of cell suspension to new culture vessels.
Incubate cultures at 37°C.
Subcultivation Ratio: A subcultivation ratio of 1:2 to 1:4 is recommended Medium Renewal: Every 2 to 3 days
Preservation:
Freeze medium: Complete growth medium supplemented with 5% (v/v) DMSO Storage temperature: liquid nitrogen vapor phase
Related Products:
recommended serum:ATCC 30-2020
References:
21937: Reddel RR, et al. Immortalized human bronchial epitherial mesothelial cell lines. US Patent 4,885,238 dated Dec 5 1989 23292: Lechner JF, et al. Asbestos-associated chromosomal changes in human mesothelial cells. Proc. Natl. Acad. Sci. USA 82: 3884-3888, 1985. PubMed: 2987952
