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The cells are positive for keratin by immunoperoxidase staining.

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.

Chen TR, et al. DLD-1 and HCT-15 cell lines derived separately from colorectal carcinomas have totally different chromosome changes but the same genetic origin. Cancer Genet. Cytogenet. 81: 103-108, 1995. PubMed: 7621404

Trainer DL, et al. Biological characterization and oncogene expression in human colorectal carcinoma cell lines. Int. J. Cancer 41: 287-296, 1988. PubMed: 3338874

Dexter DL, et al. N,N-dimethylformamide-induced alteration of cell culture characteristics and loss of tumorigenicity in cultured human colon carcinoma cells. Cancer Res. 39: 1020-1025, 1979. PubMed: 427742

Bender CM, et al. Inhibition of DNA methylation by 5-Aza-2'-deoxycytidine suppresses the growth of human tumor cell lines. Cancer Res. 58: 95-101, 1998. PubMed: 9426064

Vermeulen SJ, et al. Did the four human cancer cell lines DLD-1, HCT-15, HCT-8, and HRT-18 originate from one and the same patient?. Cancer Genet. Cytogenet. 107: 76-79, 1998. PubMed: 9809040