Different Results of IgE Binding- and Crosslinking-Based Allergy Tests Caused by Allergen Immobilization

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  • Okamoto-Uchida Yoshimi
    Division of Medicinal Safety Science, National Institute of Health Sciences
  • Nakamura Ryosuke
    Division of Medicinal Safety Science, National Institute of Health Sciences
  • Matsuzawa Yumiko
    Division of Medicinal Safety Science, National Institute of Health Sciences
  • Soma Megumi
    Division of Novel Foods and Immunochemistry, National Institute of Health Sciences Department of Food Science and Nutrition, Kyoritsu Women’s University
  • Kawakami Hiroshi
    Department of Food Science and Nutrition, Kyoritsu Women’s University
  • Ishii-Watabe Akiko
    Division of Biological Chemistry and Biologicals, National Institute of Health Sciences
  • Nishimaki-Mogami Tomoko
    Division of Novel Foods and Immunochemistry, National Institute of Health Sciences
  • Teshima Reiko
    Division of Novel Foods and Immunochemistry, National Institute of Health Sciences
  • Saito Yoshiro
    Division of Medicinal Safety Science, National Institute of Health Sciences

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<p>The physicochemical nature of allergen molecules differ from the liquid phase to the solid phase. However, conventional allergy tests are based on the detection of immunoglobulin (Ig)E binding to immobilized allergens. We recently developed an in vitro allergy testing method using a luciferase-reporting humanized rat mast cell line to detect IgE crosslinking-induced luciferase expression (EXiLE test). The aim of the present study was to evaluate the effects of antigen immobilization on the results of different in vitro allergy tests using two anti-ovalbumin (OVA) antibodies (Abs), E-C1 and E-G5, with different properties in the OVA-induced allergic reaction. Both Abs showed clear binding to OVA with an enzyme-linked immunosorbent assay and by BIAcore analysis. However, only E-C1 potentiated EXiLE response for the liquid-phase OVA. On the other hand, OVA immobilized on solid-phase induced EXiLE responses in both E-C1 Ab- and E-G5 Ab-sensitized mast cells. Western blotting of OVA indicated that E-C1 Ab binds both to OVA monomers and dimers, unlike E-G5 Ab, which probably binds only to the OVA dimer. These results suggest that antigen immobilization enhanced IgE crosslinking ability through multimerization of allergen molecules in the solid phase, resulting in an increase in false positives in IgE binding-based conventional in vitro allergy tests. These findings shed light on the physicochemical nature of antigens as an important factor for the development and evaluation of in vitro allergy tests and suggest that mast cell activation-based allergy testing with liquid-phase allergens is a promising strategy to evaluate the physiological interactions of IgE and allergens.</p>

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