The hypothesis that ribosomes are present, but may have a restricted distribution, in the Mauthner (M) axon was evaluated in isolated M-cell axoplasm after (1) staining with YOYO-1 and (2) inspection by electron spectroscopic imaging (ESI) of ribosomal RNA (rRNA) phosphorus (P). Discrete periaxoplasmic plaques, identified by their ribonuclease- sensitive fluorescence, were located circumferentially at the surface boundary of isolated axoplasm and distributed longitudinally at random intervals. Conditions that destabilized plaques, and surface blotting of plaques onto a coverslip, revealed that fluorescent puncta were probably a significant source of plaque fluorescence. Fluorescent puncta were also distributed in a delimited volume of axoplasm, subjacent to the plaque. The notably higher density of F-actin in the latter region suggested that the actin cytoskeleton may govern the spatial distribution of puncta in subcortical axoplasm. Some fluorescent plaques were superficial to the cortical F-actin layer, whereas others formed inclusions within the F-actin layer; however, plaques did not appear to contain F-actin. Periaxoplasmic plaques were also identified in ordinary myelinated axons. ESI, in which rRNA emits bright signals in the phosphorus (P) spectral line against a low- contrast background, showed that isolated axoplasm contained characteristic 25 nm P signals, which were associated or in direct contact with a pleiomorphic structural matrix, located at the surface boundary. Polyribosomal P signals were also distributed in peripheral axoplasm below the matrix. The concept of a distinct polyribosome- populated domain, distributed intermittently in the cortical zone of the axon is described. This domain is spatially defined by a plaque- like periaxoplasmic structural matrix, and a confluent volume of subcortical axoplasm integrated through an actin cytoskeleton.