Yoshimasu M A, Zhang J W, Hayakawa S, Mine Y
Department of Food Science, University of Guelph, Ont., Canada.
Int Arch Allergy Immunol. 2000 Oct;123(2):130-6. doi: 10.1159/000024431.
Buckwheat allergies are not common, however, it is considered to be a very potent allergen. Ingestion of small amounts has been found to produce anaphylactic reactions, particularly in children. Identification and characterization of the major allergen(s) in buckwheat are currently underway, however, there are some discrepancies in the findings.
Identification of the major allergen(s) was determined through Western blotting using buckwheat-allergic patients' sera. Once the allergenic proteins were identified, they were purified, their IgE-binding activity assessed through an indirect ELISA and the N-terminal amino acid sequence completed. To assess the stability of the IgE-binding epitopes, protein fractions were exposed to various treatments and assayed using an indirect ELISA. Lastly, the presence of anti-buckwheat IgG in the patients' sera was analyzed through Western blotting and ELISA.
IgE binding was detected to proteins with molecular masses of approximately 14 and 18 kDa. N-terminal sequencing was completed and found to share some homology with rice proteins associated with rice allergies and cross-allergenicity with buckwheat proteins. When the water-soluble protein fraction was heated, exposed to acidic and alkaline conditions and fully denatured, IgE-binding activity was reduced. When the fraction was partially denatured through urea, IgE-binding activity increased. Furthermore, IgG-binding activity was detected with proteins only above the 20 kDa region.
Proteins with molecular masses around 14 and 18 kDa were identified as the major allergenic proteins in the buckwheat-allergic patients' sera tested in this study. Results also indicate that these two proteins possess IgE-binding capability.
荞麦过敏并不常见，然而，它被认为是一种非常强效的过敏原。已发现摄入少量荞麦会引发过敏反应，尤其是在儿童中。目前正在对荞麦中的主要过敏原进行鉴定和表征，但研究结果存在一些差异。
通过使用荞麦过敏患者血清的蛋白质印迹法来确定主要过敏原。一旦鉴定出致敏蛋白，就对其进行纯化，通过间接酶联免疫吸附测定法评估其免疫球蛋白E（IgE）结合活性，并完成N端氨基酸序列分析。为了评估IgE结合表位的稳定性，将蛋白质组分进行各种处理，并使用间接酶联免疫吸附测定法进行检测。最后，通过蛋白质印迹法和酶联免疫吸附测定法分析患者血清中抗荞麦免疫球蛋白G（IgG）的存在情况。
检测到与分子量约为14 kDa和18 kDa的蛋白质存在IgE结合。完成了N端测序，发现其与与水稻过敏相关的水稻蛋白有一些同源性，并且与荞麦蛋白存在交叉过敏原性。当水溶性蛋白质组分被加热、暴露于酸性和碱性条件并完全变性时，IgE结合活性降低。当该组分通过尿素部分变性时，IgE结合活性增加。此外，仅在20 kDa以上区域的蛋白质检测到IgG结合活性。
在本研究中测试的荞麦过敏患者血清中，分子量约为14 kDa和18 kDa的蛋白质被鉴定为主要致敏蛋白。结果还表明这两种蛋白质具有IgE结合能力。
