黑色素
酪氨酸酶
促黑素细胞激素
体外
分子生物学
黑素细胞
细胞内
黑色素瘤
细胞培养
化学
连续稀释
原位
生物
生物化学
激素
酶
医学
癌症研究
病理
有机化学
替代医学
遗传学
作者
Walter Siegrist,Alex N. Eberlé
标识
DOI:10.1016/0003-2697(86)90327-1
摘要
A sensitive in situ melanin assay using cultured mouse B16 melanoma cells is described for structure-activity studies with melanocyte-stimulating hormone (MSH) peptides. B16 Cells were seeded at a density of 2500 cells per well in 96-well microtest tissue culture plates; after 24 h the cells were incubated in the presence of serial dilutions of MSH peptides for 3 to 5 days. The melanin released into the medium of each well was then determined spectrophotometrically at a wavelength of 405 nm using an automatic microplate reader calibrated against synthetic melanin. Studies with alpha-MSH, [Nle4, D-Phe7]-alpha-MSH, [3'-iodo-Tyr2]-alpha-MSH, adrenocorticotropin (ACTH)(1-24), and ACTH(1-39) showed that the peptides had identical intrinsic activities and that the relative potencies were similar to those obtained with a tyrosinase assay. The EC50 of alpha-MSH was 27 pM, i.e., about five- to sevenfold lower than that in the assays for tyrosinase or intracellular melanin. Thus, the new assay represents the most sensitive melanoma cell assay for MSH available to date.
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