The present paper describes the design of a HtTA (heterotetravalent allergen) as a multi-component experimental system that enables an integrative approach to study mast cell degranulation. The HtTA design allows presentation of two distinct haptens, each with a valency of 2, thereby better reflecting the complexity of natural allergens by displaying epitope heterogeneity and IgE antibody variability. Using the HtTA design, synthetic allergens HtTA-1 and HtTA-2 were synthesized to model a combination of epitope/IgE affinities. HtTA-1 presented DNP (2,4-dinitrophenyl) and dansyl haptens (Kd=22 and 54 nM for IgEDNP and IgEdansyl respectively) and HtTA-2 presented dansyl and the weak-affinity DNP-Pro (DNP-proline) haptens (Kd=550 nM for IgEDNP). Both HtTAs effectively induced degranulation when mast cells were primed with both IgEDNP and IgEdansyl antibodies. Interestingly tetravalent DNP-Pro or bivalent dansyl were insufficient in stimulating a degranulation response, illustrating the significance of valency, affinity and synergy in allergen–IgE interactions. Importantly, maximum degranulation with both HtTA-1 and HtTA-2 was observed when only 50% of the mast cell-bound IgEs were hapten-specific (25% IgEdansyl and 25% IgEDNP). Taken together, results of the present study establish the HtTA system as a physiologically relevant experimental model and demonstrates its utility in elucidating critical mechanisms of mast cell degranulation.
Design of a heterotetravalent synthetic allergen that reflects epitope heterogeneity and IgE antibody variability to study mast cell degranulation
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Michael W. Handlogten, Tanyel Kiziltepe, Basar Bilgicer; Design of a heterotetravalent synthetic allergen that reflects epitope heterogeneity and IgE antibody variability to study mast cell degranulation. Biochem J 1 January 2013; 449 (1): 91–99. doi: https://doi.org/10.1042/BJ20121088
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