Antibodies specific for protein phosphotyrosyl residues were used to localize sites of action of tyrosine-specific protein kinases in developing chick neural retina by immunoperoxidase staining. Phosphotyrosine-modified proteins became prominent in growth cone- and process-rich regions of embryonic retina during neuronal differentiation. Maximal levels accumulated in the synaptic layers and limiting membranes of the adult retina, where numerous junctional complexes reside. Two major phosphotyrosine-modified proteins in adult retina (80, 42 kDal) increased markedly during maturation. In contrast, the synaptic layers of optic tectum and other brain regions exhibited low protein phosphotyrosine levels. These results suggest a specific role for protein tyrosine phosphorylation in the retina at sites of synapses and other intercellular junctions.