Estimation of melanin content is a method for cellular melanin determination. At first, cells were seeded in a 6-well plate at a density of 2 × 105 cells and cultured for 24 h. Then, the medium was changed to conditioned medium obtained from fibroblasts cultured for 24 h. Afterward, the cell pellets were harvested and dissolved in 2.0 N NaOH, and then heated at 95 °C for 1 h. The absorbance of melanin was measured at 490 nm .