An allergen immobilization platform based on material-binding peptides for highly sensitive detection of allergies

Talanta, vol. 300, pp. 128845

Abstract

Detection of antibodies in allergy diagnostics requires allergens to be immobilized, e.g., on microtiter plates; thereby, immobilization without capturing antibodies remains a challenge. An immobilization platform, based on the material-binding peptide Snakin-1, addresses the outlined challenge for ELISA-based allergy detection on the example of Bet v 1 from birch pollen. Bet v 1 from birch pollen is the main cause of allergic rhinitis in Europe. Snakin-1 was selected among five material-binding peptides due to its polystyrene (PS)-binding properties, productive immobilization in a fusion protein with Bet v 1, and lack of interference with the primary and secondary antibody of the assay. The developed ELISA covers the whole range of antibody detection in diagnostics of the anti-Bet v 1 antibody. The reached sensitivity of the Bet v 1-Snakin-1 ELISA on standard PS plates is comparable to the Bet v 1 signal on high-binding PS plates, and the Bet v 1-Snakin-1 ELISA outperforms the Bet v 1 ELISA on high-binding plates (1.3-fold). SPR measurements of binding Bet v 1 and Bet v 1-Snakin-1 to PS, as well as subsequent antibody detection, confirm the role of Snakin-1 in productive immobilization of the Bet v 1 allergen on PS. The necessity to block the microtiter plate with BSA could be omitted, and a detection limit of at least 0.333 ng/mL (0.14 kU/L) sIgE in serum was achieved. The latter results make the scalable coating of allergens on untreated PS plates from aqueous solutions, in combination with ELISA, a highly attractive platform for allergy diagnostics. • General concept based on MBP-fusion proteins (e.g. Bet v 1-Snakin-1 fusion protein). • Identification of the potato peptide Snakin-1 as strong polystyrene binder. • Fast & oriented immobilization of allergens through fused material-binding peptides. • Highly sensitive detection of anti-Bet v 1-IgE antibodies for allergy tests.

Authors 4

1. Jana Ehlers Aachen Institute of Biotechnology

   RWTH Aachen University

   Affiliation as printed

   Institute of Biotechnology, RWTH Aachen University, Worringerweg 3, Aachen, 52074, Germany

2. Jamshid Amiri Moghaddam Aachen Institute of Biotechnology

   RWTH Aachen University

   Affiliation as printed

   Institute of Biotechnology, RWTH Aachen University, Worringerweg 3, Aachen, 52074, Germany

3. Francisca Contreras Aachen Institute of Biotechnology

   RWTH Aachen University

   Affiliation as printed

   Institute of Biotechnology, RWTH Aachen University, Worringerweg 3, Aachen, 52074, Germany

4. Ulrich Schwaneberg corresponding Aachen Institute of Biotechnology

   RWTH Aachen University

   Affiliation as printed

   Institute of Biotechnology, RWTH Aachen University, Worringerweg 3, Aachen, 52074, Germany. Electronic address: u.schwaneberg@biotec.rwth-aachen.de