Detection of allergen-induced basophil activation by expression of CD63 antigen using a tricolour flow cytometric method

In the field of allergy diagnosis, most in vitro functional tests are focus ed on basophils. Nevertheless, the very small number of circulating basophi ls limits these experiments and their clinical benefit remains controversia l. As flow cytometry is a valuable tool for identifying cell populations, e ven at low concentrations, we developed a tricolour flow cytometric method for the study of allergen-induced basophil activation. Identification of ce lls was based both on CD45 expression and on the presence of IgE on the cel l surface, since basophils express high-affinity receptors for IgE (Fc epsi lon RI). Cell activation upon allergen challenge was assessed by the expres sion of CD63 antigen on the plasma membrane. Basophil isolation and activat ion (with the chemotactic peptide formyl-methionyl-leucyl-phenylalanine) we re validated in 32 non-allergic patients. In 12 allergic patients, basophil stimulation by a relevant allergen was in most cases positive (10/12). Fur thermore a concentration-dependent hook effect was observed. Of the allergi c and non-allergic patients, none showed non-specific activation with an ir relevant allergen (specificity 100%). Overall, our preliminary results, eve n in a small population, suggest that this is a reliable and valuable metho d for the diagnosis of allergies complementing specific allergen IgE and sk in test results. Obviously, additional clinical studies are needed to valid ate these first results.