Implants from the midbrain and cerebellum of neonatal rats were cultured in roller tubes for 20 to 30 days. The cultures were incubated in a tryptamine-tetrazolium solution which demonstrated the enzyme activity of monoamine oxidase (MAO) by the formation of a formazan precipitation. Some cultures were incubated without fixation, while others were fixed in a glutaraldehyde solution prior to incubation. The specificity of the reaction was controlled by withholding the substrate or by treatment of the cultures with an MAO inhibitor. An intracellular localization of the enzyme was observed in small and medium sized neurons, but was absent in the largest neurons. Neuroglial cells, including ependyma, and mesenchymal cells and phagocytes were MAO positive. The formazan granules were scattered throughout cell bodies and into cytoplasmic processes. Glutaraldehyde fixation of cultures prior to incubation for MAO preserved the details of cell structure and enhanced the differentiation of cell types. When fixed and unfixed cultures were compared, glutaraldehyde did not appear to interfere or interact with enzyme localization; however, there was evidence that this aldehyde might be interfering with MAO concentration, since a longer incubation period for prefixed tissue was required to obtain a formazan reaction comparable to that of unfixed cultures.