Identification of a novel neuron-specific surface antigen in the developing nervous system, by monoclonal antibody...

Monoclonal antibody 4C5 was obtained after immunization of Balb/c mice with a crude membrane preparation derived from the brains of 15-day-old rat embryos. As revealed by immunocytochemistry on primary cell cultures from embryonic rat brain, it was shown that the antigen recognized by monoclonal antibody 4C5 (4C5 antigen) is localized on the cell surface of the neurons. Preliminary biochemical characterization showed that it is a peripheral protein with a molecular weight of 94,000. The 4C5 antigen does not appear to be linked with other polypeptides by S--S bonds and contains few or no disulphide intramolecular bridges. N-Glycanase digestion indicated that the protein is probably not glycosylated. Monoclonal antibody 4C5 crossreacts with membrane fractions from rat, rabbit, pig and human developing brain. It was shown by immunohistochemistry that the 4C5 antigen is widely distributed in the embryonic and adult rat brain. In the peripheral nervous system 4C5 immunoreactivity was present in dorsal root ganglion neurons. Immunoblotting and immunohistochemistry on dissociated cells from rat brain and on tissue sections of brain and dorsal root ganglia revealed an age-dependent decline in the expression of the epitope recognized by monoclonal antibody 4C5, in the central and peripheral nervous system. In particular, intense 4C5 immunoreactivity was observed during the embryonic and early postnatal ages. By the second postnatal week, expression of the protein was greatly reduced, becoming very weak at later stages of development and in the adult animal. In PC12 cell cultures, expression of the 4C5 antigen was intense in proliferating cells while being greatly reduced after nerve growth factor induced differentiation of these cells. The increased expression of the 4C5 antigen in proliferating PC12 cells and the prominent presence of this molecule during a time of neuronal migration suggest that it is involved in these developmental events.