We applied antiserum to PEP-19, a presumptive calcium-binding polypeptide, to the auditory brainstem of cats to determine whether this antiserum would selectively reveal cochlear nucleus neurons and their projections. We report that the entire populations of ventral cochlear nucleus bushy and multipolar cells, but not octopus cells, express this peptide in their somata and dendrites. Presumed axons of spherical bushy cells located dorsally and thicker globular bushy cell fibers located ventrally in the trapezoid body are immunostained, as are thin fibers presumed to represent the axons of multipolar cells. Large calyceal endings in the medial nucleus of the trapezoid body are densely immunoreactive as are smaller punctate profiles that outline immunonegative neuronal profiles in the medial and lateral superior olives. These features of immunolabeling indicate that PEP-19 is expressed in all neuronal compartments. Within the entire superior olivary complex, relatively few neurons are immunolabeled, and the vast majority of these are found in the periolivary nuclei. There are many more immunostained neurons in lateral than in medial periolivary cell groups, but their combined numbers are dwarfed by the numbers of immunolabeled cells in the ventral cochlear nucleus. The borders of the principal nuclei and some of the periolivary cell groups are well defined by the distribution of PEP-19-immunoreactive fibers and puncta. Since ventral cochlear nucleus bushy cells comprise the predominant input to principal nuclei of the superior olive, and the entire bushy cell population is immunolabeled by PEP-19 antiserum, the numbers and distribution of their inputs can be quantified. In this study we report that immunoreactive puncta apposed to the cell bodies and proximal dendrites of neurons in the medial superior olive occur at a density of 20/100 microns2. Moreover, we demonstrate by pre-embedding immunoelectron microscopy that the PEP-19-immunoreactive punctate profiles observed in the medial superior olive by light microscopy represent presynaptic terminal boutons that contain round synaptic vesicles and form asymmetric synaptic junctions, features traditionally associated with excitatory synapses. Thus, this antiserum represents a useful tool for investigating the distribution of ventral cochlear nucleus fibers and synaptic terminals within their target nuclei in the superior olive.