The isoenzymes of the glycolytic enzyme enolase have been separated and purified. The structural and functional properties of two brain enolases are described. Immunocytochemical techniques have established that one brain enolase is restricted to neuronal cells (neuron-specific enolase, NSE) while the other is localized in glial cells (nonneuronal enolase, NNE). The brain enolases, therefore, represent the first example of functional markers for neuronal and glial cell types in brain. The two enzymes are structurally distinct with the evidence establishing that they are products of separate genes. Functionally, the neuronal enolase has been demonstrated to be uniquely stable to concentrations of chloride salts that rapidly inactivate the glial enzyme. NSE may therefore represent an adaptation of this enzyme that is specifically suited to the neuronal milieu. A specific radioimmunoassay is described for NNE and NSE with the studies reported indicating that neuronal enzyme levels vary considerably when different brain areas are compared, suggesting a relationship between functional activity and levels of NSE. In addition to being a marker for neuronal cells, NSE has also been found to be present in various glands. The cells of the APUD series (amine precursor uptake and decarboxylation cells) in the pituitary, adrenal medulla, pineal, thyroid, and pancreas have been shown to contain NSE. NSE is, therefore, also a marker for these neuronlike endocrine cells since they are the only cells other than neurons that contain this protein.