Proteins S5, S6, and S54 (mol. wts. 95,000, 100,000, and 110,000 Da) appear characteristically at certain developmental stages in the chick brain (Shirao, T. and Obata, K., J. Neurochem., 44 (1985) 1210-1216). In the present study polyclonal and monoclonal antibodies were developed against electrophoretically purified S5 and S6 proteins. Each polyclonal and monoclonal antibody specifically recognized all 3 proteins, S5, S6, and S54, by immunoblot analysis. The tissue specificities of these proteins were examined by immunoblot analysis with these antibodies. Proteins S5 and S6 were found in the neural tissue and in some non-neural tissues of chick embryo. In the adult chicken, however, they were detected neither in neural nor in non-neural tissues with the exception of the spinal ganglion. Protein S54, on the other hand, was found both in late embryonic and adult neural tissues. It was detected neither in embryonic nor in adult non-neural tissues. Immunohistochemical analysis of adult nervous system showed that S54 protein was present only in neurons. Therefore it is concluded that S54 protein is a neuron-specific protein. Developmental changes of localization of these proteins were then examined by immunohistochemistry. In the developing brain, immunostaining was first observed in newly differentiated neurons, later becoming localized in the neuronal processes. In the adult brain, the immunoreactivity was mainly localized in certain types of synaptic regions, but it was also observed in a small population of neuronal somata.