Unique protein expression by the TBJ clonal derivative of C1300 murine neuroblastoma

Abstract

C1300 is a murine neuroblastoma that arose spontaneously in an A JAX mouse, and from which a clone termed TBJ was subsequently derived. C1300 is a slowly growing and poorly metastasizing tumor, whereas TBJ shows early systemic metastasis as well as aggressive local growth. Compared with TBJ cells, C1300 cells are highly immunogenic and are sensitive to natural killer cells and cytotoxic lymphocytes. In vitro, TBJ cells were found to be more rounded and less adherent than C1300 cells. Because the underlying basis for the differences between C1300 and TBJ cells has not been fully elucidated, the authors used high-resolution two-dimensional gel electrophoresis (2-DE) to study comparative aspects of total protein expression by each cell line. Of the approximately 400 individual cellular proteins that could be resolved using this technique, two were found to be reproducibly and uniquely expressed by TBJ cells and not by C1300 cells. Both proteins were anionic (pl 5.0 to 5.2) as assessed by iso-electric focusing and had molecular weights of 76,000 and 82,000 as assessed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Silver staining of SDS-polyacrylamide gels showed that the levels of 82,000-M r protein (p82) were higher than those of the 76,000-M r protein (p76). A purification protocol allowing for the isolation of p82 from TBJ cell extracts was developed, which comprised preparative two-dimensional gel electrophoresis followed by reverse-phase high-performance liquid chromatography. Full molecular identification of p82 and p76 eventually may provide new leads in the study of the metastatic or antigenic properties of neuroblastoma.