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Allergy. 2018 Jul;73(7):1425-1435. doi: 10.1111/all.13394. Epub 2018 Feb 20.

Isolation of a high-affinity Bet v 1-specific IgG-derived ScFv from a subject vaccinated with hypoallergenic Bet v 1 fragments.

Author information

1
Division of Immunopathology, Institute of Pathophysiology and Allergy Research, Center for Pathophysiology, Infectiology and Immunology, Vienna General Hospital, Medical University of Vienna, Vienna, Austria.
2
Institute of Molecular Biosciences, BioTechMed, University of Graz, Graz, Austria.
3
Division of Hematology and Hemostaseology, Department of Internal Medicine I, Vienna General Hospital, Medical University of Vienna, Vienna, Austria.
4
GE Healthcare Europe GmbH, Freiburg, Germany.
5
NRC Institute of Immunology FMBA of Russia, Moscow, Russia.

Abstract

BACKGROUND:

Recombinant hypoallergenic allergen derivatives have been used in clinical immunotherapy studies, and clinical efficacy seems to be related to the induction of blocking IgG antibodies recognizing the wild-type allergens. However, so far no treatment-induced IgG antibodies have been characterized.

OBJECTIVE:

To clone, express, and characterize IgG antibodies induced by vaccination with two hypoallergenic recombinant fragments of the major birch pollen allergen, Bet v 1 in a nonallergic subject.

METHODS:

A phage-displayed combinatorial single-chain fragment (ScFv) library was constructed from blood of the immunized subject and screened for Bet v 1-reactive antibody fragments. ScFvs were tested for specificity and cross-reactivity to native Bet v 1 and related pollen and food allergens, and epitope mapping was performed. Germline ancestor genes of the antibody were analyzed with the ImMunoGeneTics (IMGT) database. The affinity to Bet v 1 and cross-reactive allergens was determined by surface plasmon resonance measurements. The ability to inhibit patients' IgE binding to ELISA plate-bound allergens and allergen-induced basophil activation was assessed.

RESULTS:

A combinatorial ScFv library was obtained from the vaccinated donor after three injections with the Bet v 1 fragments. Despite being almost in germline configuration, ScFv (clone H3-1) reacted with high affinity to native Bet v 1 and homologous allergens, inhibited allergic patients' polyclonal IgE binding to Bet v 1, and partially suppressed allergen-induced basophil activation.

CONCLUSION:

Immunization with unfolded hypoallergenic allergen derivatives induces high-affinity antibodies even in nonallergic subjects which recognize the folded wild-type allergens and inhibit polyclonal IgE binding of allergic patients.

KEYWORDS:

Bet v 1; IgG antibody; allergy; combinatorial cloning; recombinant hypoallergenic allergen derivative

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