Use of an alpha-melanocyte-stimulating hormone analogue to improve alpha-melanocyte-stimulating hormone receptor binding assay in human melanoma.
In order to optimize the detection and measurement of alpha-melanocyte-stimulating hormone (alpha-MSH) receptivity in human melanoma cells, and the authors replaced the natural hormone by [Nle4,D-Phe7]-alpha-MSH, a more stable and potent analogue in the receptor binding assay commonly performed with alpha-MSH. The following parameters were investigated: temperature, incubation time, number of cells, and ratio of labelled to unlabelled hormone. Optimal conditions for each assay were determined. The results demonstrate that the analogue has identical binding sites to alpha-MSH, as similar reciprocal displacements of each labelled (125I) hormone by serial dilutions of unlabelled alpha-MSH or [Nle4,D-Phe7]-alpha-MSH (10(-12) M to 10(-6) M) were obtained. To further compare the two hormones, we performed a screening of various human cell lines: ten melanomas and five nonmelanomas. The assay with [Nle4,D-Phe7]-alpha-MSH yielded more receptor expression on six of ten melanoma lines against only four of ten with the natural hormone. In conclusion, the use of radiolabelled [Nle4,D-Phe7]-alpha-MSH analogue instead of labelled alpha-MSH improved both sensitivity and reproducibility in this receptor binding assay on human melanoma lines.
Libert
A
A
Laboratoire d'Oncologie et de Chirurgie Expérimentale Institut Jules Bordet, Faculty of Medicine, Université Libre de Bruxelles, Belgium.
Ghanem
G
G
Arnould
R
R
Lejeune
F J
FJ
D006801:Humans; D008545:Melanoma / Q000032:analysis / Q000378:metabolism* / Q000648:ultrastructure; D011869:Radioligand Assay / Q000379:methods; D011979:Receptors, Pituitary Hormone / Q000032:analysis / Q000378:metabolism*; D014407:Tumor Cells, Cultured / Q000032:analysis / Q000378:metabolism / Q000648:ultrastructure; D000521:alpha-MSH / Q000031:analogs & derivatives* / Q000032:analysis / Q000378:metabolism
D011979:Receptors, Pituitary Hormone; D000521:alpha-MSH; C029446:MSH receptor