Sk mel 3

Manufactured by Thermo Fisher Scientific

Sourced in France

SK-MEL-3 is a human melanoma cell line. It is a widely used model for melanoma research.

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3 protocols using sk mel 3

Cell Viability Assay with Fatty Acids

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Human foreskin fibroblast (Hs27) and human melanoma (A2058, A375, SK-Mel 3) cell lines were purchased (ATCC, Manassas VA). Cells were plated in 48-well plates at a concentration of 7.5×10³ cells/well in either DMEM (Hs27, A2058, A375; Invitrogen/GIBCO, Grand Island NY) or McCoy’s 5a Medium (SK-Mel 3; Invitrogen/GIBCO) supplemented with 10% FBS (Invitrogen/GIBCO) and incubated at 37°C in a humid ified atmosphere with 5% CO2. After 24 hours the medium was replaced with either DMEM (Hs27, A2058, A375) or McCoy’s 5a Medium (SK-Mel 3) containing 1% heat inactivated-FBS (Invitrogen/GIBCO) and 0.1% FA-free BSA (Invitrogen/GIBCO) with various concentrations of either DHA (Martek, Columbia MD), GW9508 (Cayman Chemical, Ann Arbor MI) or TAK-875 (Selleck Chemicals, Houston TX) prepared in 4μL of ETOH. DHA was provided at 25-250μM, GW9508 at 25-150μM and TAK-875 at 0-0.4μM. Controls were treated with 4μL of ETOH. All concentrations were tested in quadruplicate.
Following either 144 (DHA) or 72 hours of treatment (GW9508, TAK-875) cells were trypsinized (Trypsin-EDTA; Invitrogen/GIBCO) and viable cells were counted using an absorbance-based cell viability assay (Cell Titer Blue Cell Viability Assay; Promega, Madison WI). During the treatment, medium was changed at 72 hours for the DHA studies. Cell counts are reported as a percent of the control cell count.

Nehra D., Pan A.H., Le H.D., Fallon E.M., Carlson S.J., Kalish B.T, & Puder M. (2014). DHA, G-protein coupled receptors and melanoma: Is GPR40 a potential therapeutic target?. The Journal of surgical research, 188(2), 451-458.

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Cell Culture Protocols for Melanoma and HEK293T

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All cell lines were purchased from the American Type Culture Collection (ATCC) and maintained at 37 °C with 5% CO2. A375 (ATCC^®CRL-1619), G-361 G361 (ATCC^®CRL-1424), MeWo (ATCC^®HTB-65), SK-MEL-2 (ATCC^®HTB-68), SK-MEL-3 (ATCC^®HTB-69), SK-MEL-24 (ATCC^®HTB-71), WM2032 (ATCC^®HTB-70), and HEK293T (ATCC^®CRL-3216) cells were cultured in Dulbecco’s modified Eagle’s medium supplemented with 10% fetal bovine serum (FBS; Invitrogen), 2 mM l-glutamine, and 1% penicillin–streptomycin (Gibco-BRL). Transfections were performed according to the manufacturer’s instructions with lipofectamine 2000 (Invitrogen) or calcium phosphate transfection kit (Thermo). None of the cell lines used in this study was found in the database of commonly misidentified cell lines that is maintained by ICLAC and NCBI Biosample. All cell lines were tested and confirmed to be free of mycoplasma.

Yang Y., Luo M., Zhang K., Zhang J., Gao T., Connell D.O., Yao F., Mu C., Cai B., Shang Y, & Chen W. (2020). Nedd4 ubiquitylates VDAC2/3 to suppress erastin-induced ferroptosis in melanoma. Nature Communications, 11, 433.

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Melanoma Cell Culture and Spheroid Assay

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Human SK-MEL-3 and murine B16F10 melanoma cell lines were purchased from the American Type Culture Collection (ATCC). SK-MEL-3 cells were maintained as monolayer using culture medium consisting of McCOY’S 5A medium (Invitrogen, Cergy Pontoise, France) supplemented with 15% fetal calf serum (FCS) (Eurobio, Les Ulis, France), and 4 µg µL⁻¹ gentamycin (Invitrogen) at 37 °C in a humidified incubator containing 5% CO₂. B16F10 cells were maintained as monolayer using culture medium consisting of DMEM-Glutamax medium (Invitrogen) supplemented with 10% FCS (Eurobio), and 4 µg µL⁻¹ gentamycin (Invitrogen) at 37 °C in a humidified incubator containing 5% CO₂. The NRAS^Q61K 1007 (also named NRAS 1007) murine cell line was cultured as previously described [28 (link)]. Melanoma spheroids were generated as previously described [24 (link)].
Spheroids were harvested between 1 and 72 h post-[¹³¹I]ICF01012 removal, frozen in N₂, and stored at −80 °C during the radioactive decay (80 days; 10 times the half-life of the iodine-131 isotope) for Western blot analysis or fixed in 70% ethanol for cell-cycle studies.

Akil H., Quintana M., Raymond J.H., Billoux T., Benboubker V., Besse S., Auzeloux P., Delmas V., Petit V., Larue L., D’Incan M., Degoul F, & Rouanet J. (2021). Efficacy of Targeted Radionuclide Therapy Using [131I]ICF01012 in 3D Pigmented BRAF- and NRAS-Mutant Melanoma Models and In Vivo NRAS-Mutant Melanoma. Cancers, 13(6), 1421.

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