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Biomedical subjects

R H Butler

Publications and source records attributed to R H Butler.

24 records · Page 2Linked to original sources
PubMed1982

Antigenic sites on mouse 2.5S nerve growth factor.

Cyanogen bromide and tryptic peptides of mouse 2.5S Nerve Growth Factor (NGF) have been used to inhibit competitively binding of rabbit antiserum anti-NGF to native NGF in a solid-phase radioimmunoassay. The results showed that the larger of the two peptides obtained by cyanogen bromide cleavage (amino-acid residues 10-118) was indistinguishable from NGF in this respect, whereas the smaller N-terminal peptide (residues 1-9) was virtually non-inhibitory. Ten peptides were purified from a tryptic digest of the larger cyanogen bromide fragment. One of them, peptide G7 (residues 58-59, 60-69, 104-114, linked by disulfide bridges), was capable of almost full inhibition of binding, though only when used at a concentration about 100 times higher than that necessary to get the same effect with NGF. One other peptide, G10-H1 (residues 70-74), showed some inhibition at relatively high concentrations, but the majority of peptides, including peptide DE-5 which has been shown to be active in the NGF bioassay, were essentially non-inhibitory. A significant co-operative effect was seen when peptides G7 and G10-H1 were used in conjunction. No such effects were observed with any other combination tested.

Animals↗
PubMed1981

Monoclonal antibodies against mouse nerve growth factor produced by somatic cell hybrids.

Spleen leukocytes from rats and mice immunized against mouse 2.5S nerve growth factor (NGF) and peripheral blood leukocytes from rabbits hyper-immunized against the same antigen were fused with the mouse plasmacytoma P3X63Ag8. Hybridomas were screened by immunological assays (micro-complement fixation test and solid phase radioimmunoassay) for production of antibodies that reacted with NGF. Significant variations were seen between culture fluids from different hybrid cells. In addition, most but not all hybridoma antibodies that reacted immunologically with NGF prevented neurite outgrowth from 8-day chick embryo sensory ganglia explants after binding to NGF. These results suggest that the hybridoma antibodies produced by the different clones react with different antigenic sites on the NGF molecule.

Animals↗
PubMed1980

Nerve growth factor may stimulate either division or differentiation of cloned C1300 neuroblastoma cells in serum-free cultures.

Two clones of mouse C1300 neuroblastoma cells (clones NB1R and NB6R) bind mouse 2.5S Nerve Growth Factor (NGF) in vitro. The ligand is then capped and internalized by the cells. This step requires active cell processing. In serum-free or low serum conditions, clear effects of NGF are seen with both clones. Cultured NB1R cells are stimulated, after a lag interval of a few hours, to synthesize DNA and to proliferate, whereas NB6R cells are stimulated to cell differentiation and maintain viability under these conditions much longer than similar cultures in the absence of NGF. Stimulation of clone NB1R occurs within an optimal dose concentration of the same order as that used in the Levi-Montalcini bioassay with 8-day-old chick embryo-sensitive ganglia; the effects on clone NB6R, however, need higher NGF concentrations. Both effects are protein-specific since they are inhibited in the presence of added anti-NGF antibodies. This system could provide a convenient means to study the control of cell division in susceptible malignant neuroblastoma clones and to correlate NGF binding to receptors and biological activity.

Animals↗
PubMed1980

Protection of mice against syngeneic C1300 neuroblastoma challenge by immunization with membranes of C1300 neuroblastoma cells.

Male A/J mice 2-3 months old were inoculated sc with membranes from syngeneic C1300 neuroblastoma cells (clone NB6R) in complete Freund's adjuvant. Significant immunoprophylaxis was noted in the sensitized mice upon sc challenge with viable NB6R cells. During the experiment (60 days from viable cell challenge), each control mouse developed a palpable tumor and died within 50 days. Complete protection was obtained with a program of 4 inoculations of NB6R cell membranes. Each mouse given only 1 inoculation of NB6R cell membranes developed a palpable tumor, but afer 60 days only 1 mouse in 7 had died, which indicated a significant degree of protection. With in vitro tests of lymphocyte proliferation, rosette formation, and complement fixation, it was shown that these mice had mounted both cellular and humoral immune response against the tumor cells.

Animals↗
PubMedDate not supplied

Augmentation of the affinity of HLA class I-binding peptides lacking primary anchor residues by manipulation of the secondary anchor residues.

A direct binding assay has been used to investigate the effect of the secondary anchor residues on peptide binding to class I proteins of the major histocompatibility complex. Based on predictions from a previous chemometric approach, synthetic peptide analogues containing unnatural amino acids were synthesized and tested for B*2705 binding. Hydrophobic unnatural amino acids such as alpha-naphthyl- and cyclohexyl-alanine were found to be excellent substituents in the P3 secondary anchor position giving peptides with very high B*2705-binding affinity. The binding to B*2705 of peptides optimized for their secondary anchor residues, but lacking one of the P2 or P9 primary anchor residues was also investigated. Most such peptides did not bind, but one peptide, lacking the P2 Arg residue generally considered essential for binding to all B27 subtypes, was found to bind quite strongly. These findings demonstrate that peptide binding to class I proteins is due to a combination of all the anchor residues, which may be occupied also by unnatural amino acids-a necessary step towards the development of peptidic or non-peptidic antagonists for immunomodulation.

Amino Acid Sequence↗