Melanocytes are a specific type of epidermal cell that produce melanin in highly specialised organelles called melanosomes. Melanosomes are highly plastique, varying in shape,size and positioning within the cell. In the last years, it has become clear that “organelles dynamics”, that is the ensemble of processes that control the morphology and distribution of organelles in cells, are fundamental to shape both the cell and tissue physiology.
So far, Melanosome dynamics have been poorly studied: precise measurement of their morphology and shape has relied on classical microscopy techniques. The techniques such as transmission electron microscopy or fluorescence microscopy, which require time-consuming, manual or semi-manual detection and quantification of each individual organelle.
Here we describe a high throughput approach that we have recently developed to systematically analyse the morphology of mature melanosomes in live and fixed melanocytes: after high content imaging in brightfield, we apply a 2D image -based algorithm (Harmony™ software, Revvity) to extract the structural features of melanosomes. Specifically, we were able to quantify at single cell level the number of melanosomes as well as the area, length, width, roundness, and relative melanin content of each individual organelle.
We anticipate that this protocol will be highly relevant to both basic skin research and drug discovery. Indeed, our approach is suitable for studying the effect of different treatments and cell culture conditions on melanocyte physiology, as well as for identifying potential compounds that can be used to treat disorders associated with pigmentation defects.