Together, L10BIOBR-GFP (CRL-2770) and L10BIOBR-MAPKK (ATCC? CRL-2771) are valuable cell models for oncogenic transformation and signal transduction studies for melanoma [PubMed: 12514183].

The L10BIOBR-GFP cell line was derived as a negative control for CRL-2771. The immortalized murine melanocyte cell line L10BIOBR was transduced with pDIVA-GFP and subjected to puromycin selection.

Please note, although the L10BIOBR-GFP cell line harbors the gfp gene, as verified by PCR analysis, the cell line does not express sufficient GFP protein for detection of GFP fluorescence by flow cytometry or fluorescence microscopy.

The L10BIOBR-GFP cell line was derived as a negative control for CRL-2771. The immortalized murine melanocyte cell line L10BIOBR was transduced with pDIVA-GFP and subjected to puromycin selection. Together, L10BIOBR-GFP (CRL-2770) and L10BIOBR-MAPKK (ATCC? CRL-2771) are valuable cell models for oncogenic transformation and signal transduction studies for melanoma [PubMed: 12514183]. Please note, although the L10BIOBR-GFP cell line harbors the gfp gene, as verified by PCR analysis, the cell line does not express sufficient GFP protein for detection of GFP fluorescence by flow cytometry or fluorescence microscopy.

No, progressive tumor growth was not observed in nude mice

The L10BIOBR-GFP cell line was derived as a negative control for CRL-2771. The immortalized murine melanocyte cell line L10BIOBR was transduced with pDIVA-GFP and subjected to puromycin selection. Together, L10BIOBR-GFP (CRL-2770) and L10BIOBR-MAPKK (ATCC? CRL-2771) are valuable cell models for oncogenic transformation and signal transduction studies for melanoma [PubMed: 12514183]. Please note, although the L10BIOBR-GFP cell line harbors the gfp gene, as verified by PCR analysis, the cell line does not express sufficient GFP protein for detection of GFP fluorescence by flow cytometry or fluorescence microscopy.

Protocol:

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.
   An inoculum of 5 X 10(3) to 7 X 10(3) viable cells/sq. cm. is recommended.
6. Incubate cultures at 37°C.

Interval: Subculture when cells reach a concentration of 2 X 10(4) cells/sq. cm.

Subcultivation Ratio: A subcultivation ratio of 1:3 to 1:4 is recommended

Medium Renewal: Two to three times weekly

Freeze medium: Complete growth medium supplemented with 5% (v/v) DMSO

Storage temperature: liquid nitrogen vapor phase

Atmosphere: air, 95%; carbon dioxide (CO2), 5%

Temperature: 37.0°C