Introduction: Polcalcins belong to the family of calcium-binding proteins. They are ubiquitous in the plant kingdom and highly conserved, which leads to these panallergens showing a high degree of inter-cross-reactivity. They are responsible for allergic polysensitization, and therefore, their diagnosis is necessary for correct selection of immunotherapy. The objectives were to develop a method to purify native polcalcin with intact allergenic properties and to validate its use for diagnosis of polcalcin sensitization. Methods: Ole e 3 was purified by immunoaffinity chromatography using anti-rChe a 3 polyclonal antibodies and identified by mass spectrometry. Calcium-binding assays were performed in immunoblot and ELISA assays. Diagnostic capacity of Ole e 3 was analyzed by ELISA and compared to ImmunoCAP with sera from a pollen-sensitized population. Cross-reactivity with other polcalcins was investigated by ImmunoCAP inhibition. Results: Immunogenicity of purified Ole e 3 was not affected by the addition of calcium. However, the presence of a calcium chelator agent completely inhibited IgG binding by immunoblot and produced a 32.3% reduction in IgE binding by ELISA. Ole e 3 enabled diagnosis of polcalcin-sensitized patients, and a good correlation was revealed with ImmunoCAP. A 50% inhibition in IgE binding was obtained with 2.8 ng of Ole e 3 for rBet v 4 and 3.9 ng for rPhl p 7. Discussion/Conclusion: Native Ole e 3 was purified by maintaining its allergenic properties. This innovative method enables obtaining this active native allergen to be used for in vivo diagnosis of polcalcin sensitization.
Int Arch Allergy Immunol

中文翻译：

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Ole e 3，Polcalcin 致敏体内诊断的候选药物

简介： Polcalcins 属于钙结合蛋白家族。它们在植物界普遍存在并且高度保守，这导致这些泛变应原表现出高度的交叉反应性。他们负责过敏性多敏化，因此，他们的诊断对于正确选择免疫疗法是必要的。目的是开发一种方法来纯化具有完整致敏特性的天然 polcalcin，并验证其在诊断 polcalcin 致敏方面的用途。方法：Ole e 3 使用抗rChe a 3 多克隆抗体通过免疫亲和层析纯化，并通过质谱法鉴定。在免疫印迹和ELISA测定中进行钙结合测定。通过 ELISA 分析 Ole e 3 的诊断能力，并与来自花粉致敏群体的血清的 ImmunoCAP 进行比较。通过 ImmunoCAP 抑制研究与其他 polcalcins 的交叉反应性。结果：纯化的 Ole e 3 的免疫原性不受添加钙的影响。然而，钙螯合剂的存在通过免疫印迹完全抑制了 IgG 结合，并通过 ELISA 使 IgE 结合减少了 32.3%。Ole e 3 能够诊断对polcalcin 致敏的患者，并且与ImmunoCAP 显示出良好的相关性。使用 2.8 ng 的 rBet v 4 和 3.9 ng 的 rPhl p 7 的 Ole e 3 获得了 50% 的 IgE 结合抑制。讨论/结论：通过保持其致敏特性来纯化天然 Ole e 3。这种创新方法能够获得这种活性天然过敏原，用于 polcalcin 致敏的体内诊断。
Int Arch 过敏免疫