Epitope Mapping of Polyclonal Antibodies by Hydrogen–Deuterium Exchange Mass Spectrometry (HDX-MS)

Epitope
mapping of antibodies (Abs) is crucial for understanding
adaptive immunity, as well as studying the mode of action of therapeutic
antibodies and vaccines. Especially insights into the binding of the
entire polyclonal antibody population (pAb) raised upon vaccination
would be of unique value to vaccine development. However, very few
methods for epitope mapping can tolerate the complexity of a pAb sample.
Here we show how hydrogen–deuterium exchange mass spectrometry
(HDX-MS) can be used to map epitopes recognized by pAb samples. Our
approach involves measuring the HDX of the antigen in absence or presence
of varied amounts of pAbs, as well as dissociating additives. We apply
the HDX-MS workflow to pAbs isolated from rabbit immunized with factor
H-binding protein (fHbp), a Neisseria meningitidis vaccine antigen. We identify four immunogenic regions located on
the N- and C-terminal region of fHbp and provide insights into the
relative abundance and avidity of epitope binding Abs present in the
sample. Overall, our results show that HDX-MS can provide a unique
and relatively fast method for revealing the binding impact of the
entire set of pAbs present in blood samples after vaccination. Such
information provides a rare view into effective immunity and can guide
the design of improved vaccines against viruses or bacteria.