1a.Objectives (from AD-416)
1. Identify cross-reactive epitopes using the peptide SPOTS membrane and serum IgE from patients allergic to peanut, walnut, and almond. 2. Develop models to predict cross-reactive epitopes among the nut allergens. 3. Synthesize free peptides of the cross-reactive epitopes. 4. Determine clinically relevant cross-reactive epitopes.
1b.Approach (from AD-416)
SPOTS membrane analysis will be used to identify cross-reactive epitopes of nuts. The SPOTS data will be used to enhance the accuracy of prediction models. Free peptides of the cross-reactive epitopes will be synthesized and used to compare dissociation constants to allow the highest affinity peptides to be chosen for skin prick tests. Use SPOTS analysis, prediction tools, and inhibition analysis by basophile-histamine release analysis using basophils isolated from nuts to determine clinically relevant cross-reactive epitopes.
We utilize a computational program to predict IgE (an antibody associated with allergic responses in humans) binding sites in various tree nuts that are similar to the ones in peanuts. We then synthesize these peptides and look at actual IgE reactivity with these predicted epitopes (a region on a protein associated with allergenicity) and feed the information back into the computational model towards enhancing capabilities for prediction of cross-reactive epitopes in different nuts. This study has predicted and confirmed previously unknown epitopes or novel cross-reactive epitopes of peanut and walnut allergens.