Monoclonal antibodies against Met-enkephalin as probe in the central nervous system.

Peptides with transmitter-like properties have been found in many brain areas. Immunochemical techniques have contributed most to clarification of the function and the pathway of these substances in neuronal systems. In this paper we report the production of 4 monoclonal antibodies against Met-enkephalin and their use in studying this peptide in rat cervical cord. Two of the antibodies recognize the COOH-terminus part of the Met-enkephalin, and do not cross-react with other known peptides. The other two antibodies are mainly directed against the NH2-terminus part of the peptide. Specific interactions of these monoclonal antibodies with regions of rat cervical cord were shown by immunochemistry techniques.

Monoclonal antibodies to 5'-triphospho-(2'-5')adenyladenosine oligonucleotides.

Thirteen monoclonal antibodies to ppp(A2'p5')nA oligonucleotides have been produced by fusing Y3 myeloma cells with spleen cells of rat hyperimmunized with A2'p5'A succinyl albumin as immunogen. 125I-labeled A2'p5'A succinyl tyrosine methyl ester was used as a labeled probe. Antibodies were detected by their ability to bind the labeled analog and were selected for their affinity for A2'p5'A. There were no significant differences between the properties of the monoclonal antibodies and of the antiserum. They discriminated between 2'-5' and 3'-5' phosphodiester bonds: they crossreacted poorly with (A3'p5')2A (crossreactivity ratio, greater than 10(4)) and even less with ATP an adenosine (crossreactivity ratio, greater than (6)). The affinity was high (Kd = 6 x 10(-12) M) for succinyl A2'p5'A, which is the best ligand, and also high for A2'p5'A (crossreactivity ratio, 3) and (A2'p5')2A, (A2'p5')3A, and (As'p5')4A (crossreactivity ratio, 7.5). The binding to triphosphorylated isomers, ppp(A2'p5)nA, was affected by the presence of the triphosphate groups, and the affinity increased as the length of the isomer increased (crossreactivity ratio, 10,000 for n = 1 to 200 for n = 4). Thermodynamic analysis of these data demonstrated that, in dephosphorylated isomers, the binding site of highest affinity was located at the 5'-OH end of the molecule whereas in phosphorylated isomers, the favorable binding sites were in the middle and at the 2'-OH end of the chain. Use of monoclonal antibodies of such a specificity together with the 125I-labeled 2-5A analog allows quantification of (A2'p5')nA directly and of ppp(A2'p5')nA after removal of the terminal phosphates by alkaline phosphatase treatment.

An antigenic determinant of human beta 2-microglobulin masked by the association with HLA heavy chains at the cell surface: analysis using monoclonal antibodies.

Four anti-human beta 2-microglobulin monoclonal antibodies were produced against whole lymphoid cells or against urinary beta 2-microglobulin. Their reactivity was fully inhibited by purified soluble beta 2-microglobulin. The B1.1G6, C23.24.2, and B2.62.2 antibodies bound either to free or to HLA-complexed beta 2m. They recognized the same or very close determinants, since they achieved mutual blocking. In contrast, the C21.48A1 antibody did not bind to the cell surface and did not recognize the same determinant on the purified beta 2-microglobulin molecule as the others. It was able to bind and to form a specific complex with membrane beta 2-microglobulin only, once separated from the HLA heavy chain. These data strongly suggest that the C21.48A1 antigenic determinant might be hidden when the beta 2-microglobulin is complexed with the HLA heavy chains at the cell surface.

Production and main characteristics of a fetal calf serum-specific cell line that induces T and B cell differentiation.

Spleen cells from B6 mice injected with fetal calf serum (FCS) could be kept proliferating as a continuous cell line in vitro provided they were culture in the presence of irradiated syngeneic spleen cells and FCS. Cells in this cell line showed a strong proliferative response when stimulated with concanavalin A (Con A), and they were able to mediate the following functions: (1)they helped the generation of alloantigen-specific cytotoxic T lymphocytes (CTL) from thymocyte-spleen cell mixed lymphocyte cultures (MLC), (2)they induced the generation of CTL from normal syngeneic spleen cells in the absence of allogeneic stimulator cells, and (3)they induced normal spleen cells to differentiate into anti-sheep erythrocyte (SRBC) plaque-forming cells (PFC), in the absence of SRBC in the cultures. The use of this cell line (called line 12) may thus provide an interesting approach for the study of cellular and molecular requirements for cell-cell interactions and for the differentiation of T and B effector functions.

Growth characteristics in vitro of hybrids between normal and transformed cell lines.

Sensitivity to the inhibition of division during cell crowding in vitro was determined for hybrids between normal and tumor cell populations of human and mouse origin. Interspecies hybrids were more sensitive to cell crowding than intraspecies hybrids. The results suggest that the inhibition of cell division due to crowding was more dependent upon the species of origin rather than on the normal or transformed character of the parental cell lines of the hybrid cell populations.