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Monoclonal antibodies to the dentate gyrus: immunocytochemical characterization and flow cytometric analysis of hippocampal neurons bearing a unique cell-surface antigen

Monoclonal antibodies were generated using 5 d neonatal rat dentate gyrus as immunogen. One antibody of this panel, G6E3, recognized a cell- surface protein with an Mr = 43,000 that was found only in the nervous system. The antigen was expressed as early as embryonic day 13 in the rat in both the brain and spinal cord. In the adult rat the antigen was demonstrated immunohistochemically to be restricted to dentate gyrus granule, hippocampal pyramidal, and cerebellar Purkinje neurons. These results suggested that the antigen recognized by G6E3 may be developmentally regulated. Moreover, G6E3 did not appear to bind to mitotic cells, implying that the antigen was expressed after the terminal mitosis. The antibody also bound to hippocampal and cerebellar cells from mouse brain, including the reeler mutant, and rat hippocampal neurons in vitro. Double-labeling experiments performed on embryonic rat hippocampal cultures with G6E3 and antibodies to neuron- specific enolase (NSE) or anti-glutamic acid decarboxylase (GAD) revealed that only NSE-positive cells were immunoreactive for G6E3 and, while G6E3-positive cells were decorated with GAD-positive boutons, their cell bodies did not contain GAD. With the use of a fluorescence- activated cell sorter it was possible to analyze the immune reaction on embryonic and postnatal hippocampal cells and to sort G6E3-positive neurons for maintenance in vitro.