Transmembrane control of the receptors on normal and tumor cells: I. Cytoplasmic influence over cell surface components☆

Consequences of blocked sterol synthesis in cultured cells. DNA synthesis and membrane composition.

Hybridoma cells have been derived from a fusion between mouse myeloma cells (P3-NSI/1 Ag4) and spleen cells from a mouse immunized with membrane-enriched fractions from the human melanoma cell line Me-43. Of the 26 hybrids obtained, seven secreted antibodies which reacted with the melanoma cell line used for immunization as assessed by an indirect antibody-binding radioimmunoassay. The specificity of the antibodies produced by the seven positive hybrids was further investigated on 16 melanoma cell lines, 15 other tumors, and 14 lymphoblastoid cell lines. The antibodies from four positive hybrids showed a broad reactivity, whereas those from three hybrids reacted exclusively with melanoma cells. The antibodies from two of these three hybrids, α-Mel/5 and α-Mel/14, seem to be directed against common melanoma antigen(s) since they reacted with all (with one exception) of the 16 melanoma cell lines tested, whereas antibodies from the third hybrid, α-Mel/7, reacted only with five of the 16 melanoma lines. Reciprocal binding inhibition tests using [3H]leucine-labeled antibodies showed that α-Mel/5 and α-Mel/14 antibodies were directed against different antigenic determinants.

Common human melanoma-associated antigen(s) detected by monoclonal antibodies.

Spectrin: current understanding of its physical, biochemical, and functional properties.

In order to investigate the properties of the membrane-bound IgE-receptor complex, a simple procedure has been adapted for preparing large plasma membrane vesicles from rat basophilic leukemia cells. These vesicles pinch off from the adherent cells after treatment with 2 mM N-ethylmaleimide or 50 mM formaldehyde and 1 mM dithiothreitol, and they are isolated from the supernatant after two centrifugation steps with yields of 20-25% of the initial cell-bound 125I-IgE. With phase and fluorescence microscopy, micron-size vesicles are seen which are unilamellar and spherically shaped and devoid of intracellular organelles. On dextran gradients at least 70% of the 125I-IgE is bound to membranes which band at low density, indicating large, intact vesicles that are impermeable to macromolecules. Between 60 and 75% of the bound 125I-IgE is accessible to the external medium, showing the vesicles to be predominantly right side out. This preparation was found to be suitable for resonance energy-transfer measurements. We have determined that amphipathic, fluorescent donor and acceptor probes partition into the vesicle bilayer in a randomly distributed, noninteracting manner. The densities of the probes can be ascertained directly from the amount of energy transfer that is observed as a function of acceptor concentration. This experimental system will allow energy-transfer measurements to determine distances between sites on receptor-bound IgE and the membrane surface.

Structural studies on the membrane-bound immunoglobulin E-receptor complex. 1. Characterization of large plasma membrane vesicles from rat basophilic leukemia cells and insertion of amphipathic fluorescent probes

Analysis of the proteins in thymocyte plasma membrane and smooth endoplasmic reticulum by sodium dodecylsulfate-gel electrophoresis

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Analysis of the proteins in thymocyte plasma membrane and smooth endoplasmic reticulum by sodium dodecylsulfate-gel electrophoresis