Recognition of a monoclonal antibody against a small molecular weight antigen by monitoring the antigen–antibody reaction using fluorescence labeled structured peptides

Interaction between proteins (as analytes) and de novo designed structured peptides as capture molecules cause structural changes, which are reflected in fluorescent-intensity changes of labeled peptides in a dose dependent manner. In contrast to conventional detection methods our detection system does not involve the detection of specific molecules themselves in a 1:1 manner, but uses the principle of the differences in fluorescent intensity changes of capture peptides upon addition of analytes. Instead of the use of secondary antibodies we have attempted monitoring these structural changes by an array of de novo designed synthetic and structured peptides. In the present study we have focused on a recognition system, 5-fluorouracil, as a low molecular antigen and a monoclonal antibody against 5-FU. The fluorescent intensity changes of fluorescent labeled peptides have been measured after incubation with a monoclonal antibody and again after further incubation with the antigen, 5-FU. Unique intensity changes were found for several peptides in the fluorescent peptide library that allowed the visualization as a color-coded protein fingerprint. The peptide screen used in the present study offers a useful detection system as capture molecules for peptide-based microarrays.

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