Increase in CD63 Expression in Amoxicillin Allergic Patients Upon Basophil in Vitro Stimulation with LPS and Amoxicillin in Immediate Reactions

The basophil activation test (BAT) based on CD63 or CD203c expression is a currently accepted tool to help in the diagnosis of IgE-mediated reactions to betalactams. However, it has not optimal sensitivity. One factor could be related to pathogen-derived molecules that, although absent in the test tube, could play a role in the acute phase of allergic reactions. In addition, the mechanism of induction of CD203c and CD63 during basophil activation can differ. We hypothesize that adding these molecules to the BAT would increase sensitivity, but they may act differently on CD63 versus CD203c detection. Samples from twenty-one allergic patients (6 urticaria, 11 anaphylaxis, 3 anaphylactic shock) and 20 tolerant subjects were assessed using amoxicillin (AX) alone (conventional BAT), or in combination with different Toll-like receptor (TLR) 1-9 ligands. The percentage of CD63 and CD203c expression was compared, and analytical parameters were calculated. Among all TLR ligands, LPS, TLR4 ligand, was the most potent BAT enhancing factor. Percentage of CD63 raised significantly after LPS+AX stimulation compared to AX or LPS stimulation alone, only in allergic patients. Sensitivity of CD63-LPS-BAT raised from 42% to 67% compared to conventional BAT with a specificity of 85%. Importantly, 50% of undetected patients by conventional BAT were recovered. In contrast, neither sensitivity nor specificity of CD203c-BAT were increased by the addition of LPS. This study highlights the differences between CD203c and CD63 molecules in basophil´s response to pathogen receptor stimulation and points out the diagnostic potential of LPS as a BAT cofactor.