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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 pipetting gently.
5. Add appropriate aliquots of cell suspension to new culture vessels. An split ratio of 1:5 to 1:10 is recommended.
6. Incubate culture vessels at 37°C.

Hai M, et al. Comparative analysis of Schwann cell lines as model systems for myelin gene transcription studies. J. Neurosci. Res. 69: 497-508, 2002. PubMed: 12210843

Bansal R, Pfeiffer SE. Regulated galactolipid synthesis and cell surface expression in Schwann cell line D6P2T. J. Neurochem. 49: 1902-1911, 1987. PubMed: 2824698