蚊虫唾液过敏原埃及伊蚊Aed a 3：克隆、全面分子分析及临床评估
Mosquito salivary allergen Aed a 3: cloning, comprehensive molecular analysis, and clinical evaluation.
作者信息
Peng Z, Xu W W, Sham Y, Lam H, Sun D, Cheng L, Rasic N F, Guan Q, James A A, Simons F E R
机构信息
Department of Pediatrics and Child Health, University of Manitoba, Winnipeg, MB, Canada.
Department of Immunology, University of Manitoba, Winnipeg, MB, Canada.
出版信息
Allergy. 2016 May;71(5):621-8. doi: 10.1111/all.12812. Epub 2016 Mar 6.
BACKGROUND
Allergic reactions to mosquito bites are an increasing clinical concern. Due to the lack of availability of mosquito salivary allergens, they are underdiagnosed. Here, we reported a newly cloned mosquito Aedes (Ae.) aegypti salivary allergen.
METHODS
A cDNA encoding a 30-kDa Ae. aegypti salivary protein, designated Aed a 3, was isolated from an expression library. The full-length cDNA was cloned into a baculovirus expression vector, and recombinant Aed a 3 (rAed a 3) was expressed, purified, and characterized. Skin prick tests with purified rAed a 3 and Ae. aegypti bite tests were performed in 43 volunteers. Serum rAed a 3-specific IgE levels were measured in 28 volunteers.
RESULTS
The primary nucleotide sequence, deduced amino acid sequence, and IgE-binding sites of Aed a 3 were identified. rAed a 3-selected antibodies recognized a 30-kDa Ae. aegypti saliva protein. rAed a 3 bound IgE in mosquito-allergic volunteers and the binding could be inhibited by the addition of natural mosquito extract dose dependently. Immediate skin test reactions to rAed a 3 correlated significantly with mosquito bite-induced reactions. Of the bite test-positive volunteers, 32% had a positive rAed a 3 skin test and 46% had specific IgE. No bite test-negative volunteers reacted to rAed a 3 in either the skin tests or the IgE assays, confirming the specificity of the assay.
CONCLUSIONS
Aed a 3 that corresponds to the Aegyptin protein is a major mosquito salivary allergen. Its recombinant form has biological activity and is suitable for use in skin tests and specific IgE assays in mosquito-allergic individuals.
背景
蚊虫叮咬引起的过敏反应在临床上越来越受到关注。由于缺乏蚊虫唾液过敏原，此类过敏反应常常被漏诊。在此，我们报告了一种新克隆的埃及伊蚊唾液过敏原。
方法
从一个表达文库中分离出编码一种30 kDa埃及伊蚊唾液蛋白（命名为Aed a 3）的cDNA。将全长cDNA克隆到杆状病毒表达载体中，表达、纯化并鉴定重组Aed a 3（rAed a 3）。对43名志愿者进行了纯化的rAed a 3皮肤点刺试验和埃及伊蚊叮咬试验。对28名志愿者检测了血清中rAed a 3特异性IgE水平。
结果
确定了Aed a 3的初级核苷酸序列、推导的氨基酸序列和IgE结合位点。rAed a 3筛选出的抗体识别一种30 kDa的埃及伊蚊唾液蛋白。rAed a 3能与蚊虫过敏志愿者的IgE结合，加入天然蚊虫提取物后，这种结合可被剂量依赖性抑制。对rAed a 3的即刻皮肤试验反应与蚊虫叮咬引起的反应显著相关。在叮咬试验阳性的志愿者中，32%的人rAed a 3皮肤试验呈阳性，46%的人有特异性IgE。在皮肤试验或IgE检测中，叮咬试验阴性的志愿者对rAed a 3均无反应，证实了该检测方法的特异性。
结论
与埃及伊蚊蛋白相对应的Aed a 3是一种主要的蚊虫唾液过敏原。其重组形式具有生物活性，适用于蚊虫过敏个体的皮肤试验和特异性IgE检测。
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