An antiserum against the PC12 cell line defines cell surface antigens specific for neurons and Schwann cells

Antibodies raised in rabbits and in guinea pigs against nerve growth factor-treated PC12 cells were absorbed exhaustively with three non-neuronal cell lines. In immunofluorescent staining experiments, these absorbed sera, designated anti-PC12, labeled specifically several different types of cultured neurons, including cerebral, cerebellar, spinal, dorsal root, and superior cervical neurons. Non-neuronal cells in these cultures, such as astrocytes, oligodendrocytes, and fibroblasts, were not labeled. However, neural crest-derived adrenal chromafhn cells and Schwann cells were stained by anti-PC12. Further absorption with adrenal tissue rendered anti-PC12 specific for neurons and Schwann cells, while still further absorption with Schwann cells yielded an antiserum that was reactive only with dorsal root and superior cervical neurons. Absorption of anti-PC12 with adult brain resulted in the loss of activity against all cells except PC12 cells. These absorption experiments suggested that anti-PC12 recognized at least four distinct cell surface components. Some anti-PC12-reactive components were identified by using the anti-PC12 serum to prepare immune precipitates from detergent extracts of 12”1-labeled cells. The immune precipitates were analyzed by polyacrylamide gel electrophoresis. Anti-PC12 precipitated two components with apparent molecular weights of 190,000 and 140,000 that were common to Schwann cells, chromaffin cells, and all types of neurons. Smaller quantities of these two components could also be precipitated from cerebellar glial cells. In addition, anti-PC12 precipitated a glycoprotein of greater than 200,000 daltons from the cultured neurons, the neuronal cell lines, and the Schwann cells, but not from chromaffin cells or from cerebellar glia. Each neuronal cell type expressed a characteristic form of this large glycoprotein, as judged by electrophoretic mobility. For example, the molecules found on four cell lines, PC12, B35, N18, and /3HC, had apparent molecular weights of 235,000, 225,000, 220,000, and 215,000, respectively. The 235,000-dalton glycoprotein from PC12 cells is likely to be identical to the NILE glycoprotein described by McGuire et al. (McGuire, J., L. Greene, and A. Furano (1978) Cell 15: 357-365), while the 220,000-dalton component from N18 cells is probably similar to that described by Akeson and Hsu (Akeson, R., and W. Hsu (1978) Exp. Cell Res. 115: 367-377). Adrenal-absorbed anti-PC12 did not precipitate the 190,000and 140,000-dalton components from any of the cell types but continued to precipitate the 215,000to 235,000-dalton glycoproteins from the neurons and Schwann cells. These high molecular weight glycoproteins thus appear to be cell surface markers for neurons and Schwann cells. The PC12 cell line (Greene and Tischler, 1976) shares many properties with sympathetic neurons and has thus provided an extremely useful model system for studying several types of neuronal mechanisms, including the function of neuronal acetylcholine receptors (Patrick and ’ This work was supported by a Muscular Dystrophy Association research grant, by National Institutes of Health Grant NS 16112 to W. B. S., and by a Muscular Dystrophy Association postdoctoral fellowship to J. M. L. We thank Dr. David Schubert and Dr. Simon Halegoua for their helpful comments concerning these experiments. ‘To whom correspondence should be addressed at Neurobiology Laboratory, The Salk Institute, P.O. Box 85800, San Diego, CA 92138. Stallcup, 1977a, b; Stallcup and Patrick, 1980; Dichter et al., 1977), Na+ channels (Stallcup, 1979; Dichter et al., 1977), Ca2+ channels (Stallcup, 1979; Ritchie, 1979), K’ channels (Arner and Stallcup, 1981), the synthesis and secretion of acetylcholine and catecholamines (Ritchie, 1979; Schubert et al., 1977; Greene and Rein, 1977), and the response to nerve growth factor (McGuire et al., 1978; Schubert et al., 1978). In light of the many similarities between PC12 cells and neurons, it seems likely that antisera raised against PC12 cells might define cell surface antigens specific to neurons. Lee et al. (1977, 1981) have shown that antisera raised against PC12 cells and absorbed with liver, kidney, spleen, and thymus do con-