RGB method in immunofluorescence investigations on stem cells

Colour is not related to a particular discipline, but it is transversely present in many circles and inalmost all the aspects of life. It has a special value in art, but also as far as other disciplines areconcerned, like the sciences, the colour is at the basis of some of their intrinsic significances and it oftenneeded to allow the interpretation of some of their phenomena as well. As regards the development ofcell biology knowledge, colour acquired more and more importance in revealing the observations of theresearchers. A field in which the methods based on the colours are particularly employed is theimmunofluorescence, used to identify specific proteins in cells and tissues. These techniques combinethe fluorochrome properties with specific molecules, i.e. antibodies, directed against particularsubstances to investigate, for example a specific protein. In single immunofluorescence analysis, thesignal from an excited fluorochrome corresponds to a particular protein. In multiple immunofluorescenceanalysis, two or more signals are simultaneously detected to show the localization of differentproteins on the same sample. The three primary colours red, green and blue were currently assigned tothe signals from immunofluorescence-processed samples and visualized by the RGB method. In thepresent work, different examples of RGB applications in immunocytochemical investigations areshowed: the first concerns the multiple analysis of three markers, localized in different loci of the cellplasma membrane; the second is related to the co-localization of two signals in the same site of specificsubcellular structures. In this case the secondary colours, obtained by overlapping the primary ones,demonstrate the specific co-presence of two proteins in the same site. With the present paper, theauthors wish to underline the relevant role of colours also in those areas in which colours are the meansnot the end.