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The sNF94.3 cell line was derived from a lung metastasis diagnosed by histopathology as MPNST (Malignant Peripheral Nerve Sheath Tumor) in a patient meeting NF1 diagnostic criteria. The cells were derived from numerous passages of primary tumor material in culture until they were a homogenous Schwann cell-like population which displayed a clonal morphology immunopositive for both the cytoplasmic Schwann cell markers S100 and p75.

female

43 years

This cell line has a normal karyotype. Western immunoblotting showed apparently full-length neurofibromin protein.

Volumes used in this protocol are for 75 cm² flasks; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes.

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 which 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. Transfer cell suspension to a centrifuge tube and spin at approximately 125 X g for 5 to 10 minutes. Discard supernatant.
6. Resuspend the cell pellet in fresh growth medium. Add appropriate aliquots of the cell suspension to new culture vessels. An inoculum of 5 X 10³ to 7 X 10³ viable cells/cm² is recommended.
7. Incubate cultures at 37°C. Subculture when cell concentration is between 8 X 10⁴ and 1 X 10⁵ cells/cm²

Subcultivation Ratio: A subcultivation ratio of 1:4 to 1:6 twice weekly is recommended

Freeze medium: Culture medium,90%; DMSO,10%

Storage temperature: Liquid nitrogen vapor phase

Temperature: 37°C

Amelogenin: X

CSF1PO: 11,12

D13S317: 11,12

D16S539: 9,12

D5S818: 10,12

D7S820: 9,14

THO1: 7,9.3

TPOX: 11,12

vWA: 17,18

The cells were derived from numerous passages of primary tumor material in culture until they were a homogenous Schwann cell-like population which displayed a clonal morphology immunopositive for both the cytoplasmic Schwann cell markers S100 and p75.