FIGURE 1. Detection of allergen sensitization by quantifying
binding of fluorescent allergen components of BV and RGP to basophils.
A) Schematic diagram of DNA constructs for generation of the
recombinant allergens Api m 1, Lol p 1 and Lol p 5. Recombinant
allergens contained mutations to render the catalytic site inactive (Api
m 1 H67Q, Lol p 1 H104V). Leader peptide and spacer (L); 6-histidine tag
(His); avidin tag (AviTag). B) Western blots of recombinant
allergens detected with anti-His antibody. C) Representative
flow plot for detection of circulating basophils
(CD123+IgE+) within gated
mononuclear cells (Suppl Fig 1 ). D) Histograms
depicting representative fluorescent staining of Api m 1, Lol p 1 and
Lol p 5 tetramers on basophils from relevant allergic patients (red) and
controls (blue). Values above the histograms represent median
fluorescence intensity values of the population.