In agreement with its distinct phylogenetic origin, the envelope of Thermus thermophilus consists of a complex pattern of layers with properties intermediate between those of Gram positives and Proteobacteria. Its cell wall of Gram positive composition is surrounded by an outer envelope that includes a crystalline layer scaffold built up by the SlpA protein, lipids and polysaccharides. The synthesis of this outer envelope has been studied by confocal microscopy. Available amino groups from the cell surface, mainly belonging to the SlpA protein, were covalently labelled in vivo with fluorescent dyes. Stained cells were able to grow without any apparent loss of viability, allowing the localization of the regions of new synthesis as dark nonfluorescent spots. Our results demonstrate that the outer envelope of T. thermophilus is synthesized from a central point in the cells, likely following a helical pattern. Cell poles and subpolar regions are basically inert and retain their label for generations.