Chemical shift differences between free and Fab-bound peptide correlate with a two-stage selection of peptide sequences from a random phage display library to delineate critical and non-critical residues for antibody recognition

Epitope libraries provide a method to identify peptide ligands for antibodies, receptors or other binding proteins. As such, they provide a powerful tool to rapidly identify lead ligands in the drug discovery process. In an attempt to correlate structural information with the results from peptide screening, we have used NMR spectroscopy of peptide/antibody complexes to demonstrate that core residues identified through a two-stage selection process undergo a larger structural change upon binding antibody than do positions in the peptide amenable to a variety of side chains. The model system used was the M2 monoclonal antibody/Flag? octapeptide epitope system. We have analyzed two peptides: Ac-Asp-Tyr-Lys-Leu-Gly-Asp-Asp-Leu-NH₂ (peptide l), which contains several non-core positions randomized, and Ac-Asp-Tyr-Lys-Asp-Asp-Asp-Asp-Leu-NH₂ (peptide 2), which closely corresponds to the original Flag? sequence. Enrichment of the peptides with ¹⁵N facilitated the investigation by permitting spectral editing of the peptide resonances in the presence of antibody. For peptide 1 the absolute shifts for the free vs. Fab-bound peptide were found to be largest for the amide groups of Asp-1 and Asp-6, in agreement with classification of these residues as critical by the phage display library selection process. For peptide 2 the largest absolute shifts were observed for Asp-1 and Asp-4, with the other aspartic acid residues also showing significant but smaller changes. © Munksgaard 1995.     

Immunoblotting of streptococcal antigens in guttate psoriasis

Guttate psoriasis may be precipitated by acute streptococcal infection, usually of the upper respiratory tract. We have studied the immune response to streptococci in 26 patients presenting with a first or recurrent episode of acute guttate psoriasis (AGP), using immunoblotting. Eighteen of 26 patients studied had a demonstrable response to a wide range of streptococcal antigens using this approach, compared with 14 of 26 patients who demonstrated a response using more conventional anti-streptococcal antibody tests. Patients with AGP had a significantly higher antibody detection score using immunoblotting than did control subjects (P<0.005). We conclude that immunoblotting is a useful technique in studying this condition and may be of benefit in exploring the immunopathogene-sis of AGP.     

Inhaled fentanyl as a method of analgesia

A study was undertaken to investigate the use of fentanyl by aerosol for postoperative analgesia. Seven patients had placebo, six received fentanyl 100 micrograms and seven were given fentanyl 300 micrograms. A significant improvement in postoperative pain, as assessed by linear visual analogue scale, was achieved in the higher dose group, and in both fentanyl groups the time to alternative analgesia was significantly longer than in the control group. Serum fentanyl levels after inhalation of 100 micrograms reached a plateau around 0.04 ng/ml and after 300 micrograms at around 0.1 ng/ml after 15 minutes. Inhaled fentanyl may have a useful analgesic effect despite these low serum levels; this supports the hypothesis that the mode of analgesia from inhaled opioids may be different from that after other routes of administration. There were no adverse effects such as respiratory depression, bronchospasm, nausea or drowsiness.