APPLICATION OF THE NEW STEREOLOGICAL PROBES TO THE STUDY OF THE MELANOSOME IN CLOUDMAN S91 MELANOMA-CELLS

The relationship between melanosome size and number and melanin conten t has been investigated in Cloudman S91 melanoma cells growing in vitr o using both ''model-based'' and ''design-based'' stereological proced ures. Cells were cultured for 4 days, harvested at daily intervals, an d resin-embedded for light and electron microscopy; one aliquot of eac h sample of cells was assayed to determine its melanin content. By com paring their volume-weighted mean nuclear volume and their number-weig hted mean nuclear volume, we have found that the nuclei of Cloudman me lanoma cells form a fairly homogeneous population. The volume fraction and absolute volume of premelanosomes (V-Vpm,V-cell and V-pm) and mat ure melanosomes (V-Vmm,V-cell and V-mm) were all found to decrease pro gressively throughout the period of culture as did the number of preme lanosomes (N-pm) and mature melanosomes (N-mm). Whilst the volume-weig hted mean volume of individual stage I and stage II premelanosomes, (( V) over bar(Vipm)), remained fairly constant at about 10 nm(3), the vo lume of individual stage III and IV mature melanosomes showed signific ant variation ranging between about 13 nm(3) and 32 nm(3). The melanin content of the cells decreased progressively over the 4 days of cultu re. There were, however, considerable variations in both the average m elanin content per unit volume of mature melanosomes, in the range 170 -600 fg/mu m(3), and in the melanin content per individual mature mela nosome, in the range 3-12 fg. Our findings show that stereological tec hniques can provide unbiased and sensitive tools for the study of the morphological basis of melanogenesis; their value will become even mor e evident when they are combined with techniques for the localization of melanogenic enzymes and their substrates.