Introduction
Current food allergy diagnostics comprise careful history, skin prick
tests, measuring specific IgE (sIgE) and double-blind placebo-controlled
food challenges (DBPCFC) as the gold standard. However, DBPCFCs are
burdensome for the patient, costly and require dedicated hospital
facilities1, 2. On the other hand, current
technologies to measure sIgE detect both clinically relevant and
irrelevant sensitization, potentially leading to incorrect diagnosis and
consequently unnecessary food restrictions3, 4.
Clinically relevant peanut sensitization is associated with sIgE against
the major peanut allergens belonging to the 2S albumin family, Ara h2
and 6. In previous studies in adults, 100% positive predictive values
for sIgE against Ara h2 and 6 were found using sIgE positivity
thresholds of respectively 1.75 kU/l and 1.8 kU/l. Specific IgE levels
below these thresholds, however, overlapped between allergic and
tolerant subjects, preventing precise diagnosis5, 6.
The occurrence of clinically irrelevant sensitization to Ara h2 and 6
might be explained by differences in peanut (Ara h2 and 6) specific
antibody repertoires. These differences may include the presence of
non-IgE antibodies blocking the binding of clinically relevant epitopes
by sIgE of tolerant patients. Moreover, differences may also be based on
antibody affinity and/or epitope recognition patterns. So far, no clear
differences between allergic and tolerant subjects were observed by
epitope mapping approaches7. These evaluations may
have been hampered by the use of patient sera consisting of polyclonal
IgE antibodies. Sera of allergic subjects usually contain mixtures of
antibodies recognizing both clinically relevant and irrelevant epitopes.
On the other hand, sera from tolerant subjects might contain antibodies
recognizing clinically relevant epitopes with insufficient affinity for
successful FcεRI receptor crosslinking, as well as antibodies
recognizing clinically irrelevant epitopes8. Hence,
deep analysis of monoclonal antibodies (mAbs) from specific B-cells may
provide more insights into differences in specific antibody repertoires
between allergic and tolerant subjects.
To this end, we analyzed gene sequences encoding the variable region of
peanut 2S albumin-specific mAbs from 6 allergic and 6 sensitized but
tolerant adults. In particular, sequence motifs of the HCDR3 region, the
most important region for recognizing antigens9, were
hierarchically clustered. Clustering of HCDR3 regions resulted in four
motifs exclusively present in allergic donors and three motifs
associated with tolerance.