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Food Science and human Wellness 2023年第3期12卷 755-764页

作者： Minjia Wang Shuo Wang Xiaodong Sun Zhirui Deng Bing Niu Qin Chen School of Life Sciences Shanghai UniversityShanghai 200444China Medical School Shanghai UniversityShanghai 200444China

Little information was so far available about allergenic mechanism of the roasted peanut allergens during initial stages of allergy.The purpose of this study was to determine the influence of roasting(150℃,20 min)on biochemical and biological properties of Ara h 3,a major peanut allergen.Allergenicity of roasted peanut emulsion to mice,differences in uptakes between Ara h 3 purified from raw peanuts(named as Ara h 3-Raw)and that purified from roasted peanuts(named as Ara h 3-Roasted)by bone marrow-derived dendritic cells(BMDCs)and the implication of cell surface receptors involving in uptake,and changes in glycosylation and structure of Ara h 3 after roasting were analyzed in this study.This study suggested that roasting increased allergenicity of peanut to BALB/c mice.Maillard reaction and structural changes of Ara h 3 induced by roasting significantly altered the uptake of Ara h 3-Roasted by BMDCs,and modified Ara h 3 fate in processes involved in immunogenicity and hyper allergenicity,indicating that food processing pattern can change food allergenicity.

关键词： Roasted peanut Ara h 3 Bone marrow-derived dendritic cells(BMDCs) Allergenicity Maillard reactions

湿热处理对花生球蛋白Ara h 3抗原性的影响

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河南工业大学学报（自然科学版） 2023年第6期44卷 44-50页

作者：吴枭席俊孙富宇杨欢欢水天娇黄芃博高一达河南工业大学粮油食品学院河南郑州450001

为探究湿热处理对花生球蛋白Ara h 3抗原性的影响,采用硫酸铵分级沉淀法从花生脱脂粉中分离纯化出Ara h 3,进行不同温度、时间和质量浓度的湿热处理,通过正交试验确定最优湿热处理条件,采用间接ELISA法检测湿热处理后Ara h 3抗原性变化... 详细信息

为探究湿热处理对花生球蛋白Ara h 3抗原性的影响,采用硫酸铵分级沉淀法从花生脱脂粉中分离纯化出Ara h 3,进行不同温度、时间和质量浓度的湿热处理,通过正交试验确定最优湿热处理条件,采用间接ELISA法检测湿热处理后Ara h 3抗原性变化,测定外源荧光、游离巯基含量以及圆二色光谱,从蛋白结构的变化解释湿热处理对Ara h 3抗原性的影响。研究表明:湿热处理能够降低Ara h 3抗原性,且在质量浓度10 mg/mL、90℃处理30 min时,抗原性降至最低;湿热处理后的Ara h 3荧光强度、表面疏水性增强,三级结构发生变化;游离巯基含量增加、空间结构发生改变;α-螺旋和β-折叠含量降低,无规则卷曲含量增加,二级结构发生变化。湿热处理后Ara h 3空间结构发生改变,部分抗原表位遭到破坏或被包埋,抗原性降低。

关键词：湿热处理 Ara h 3 抗原性蛋白结构酶联免疫吸附法

花生致敏蛋白Ara h 3单克隆抗体的制备及其免疫学特性鉴定

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粮食与油脂 2023年第9期36卷 79-83页

作者：孙富宇席俊范雨函水天娇杨欢欢尚阿晨河南工业大学粮油食品学院河南郑州450001

以Ara h 3为免疫原免疫小鼠。选取血清效价高的小鼠再次免疫,制备出脾细胞后与骨髓瘤细胞一起进行细胞融合,得到杂交瘤细胞株,筛选阳性株制备大量单克隆抗体。通过酶联免疫吸附试验(ELISA)对制备的单克隆抗体进行效价测定、敏感性测定... 详细信息

以Ara h 3为免疫原免疫小鼠。选取血清效价高的小鼠再次免疫,制备出脾细胞后与骨髓瘤细胞一起进行细胞融合,得到杂交瘤细胞株,筛选阳性株制备大量单克隆抗体。通过酶联免疫吸附试验(ELISA)对制备的单克隆抗体进行效价测定、敏感性测定和特异性测定。通过蛋白质免疫印迹试验进一步验证其特异性。结果表明:免疫性较好的4B2单克隆抗体经纯化后的效价为1∶5.12×10^(5)。由敏感性测定得知4B2单克隆抗体的半抑制浓度(IC_(50))为389 ng/mL,证明其具有较好的敏感性,交叉反应率小于0.5%,成功制备出纯度高、效价高、敏感性好、特异性强的Ara h 3鼠源单克隆抗体,为建立免疫学快速检测方法奠定了基础。

关键词： Ara h 3 单克隆抗体酶联免疫吸附试验

Selection of specific nanobodies against peanut allergen through unbiased immunization strategy and the developed immuno-assay

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Food Science and human Wellness 2023年第3期12卷 745-754页

作者： Yaozhong hu Chuan Zhang Jing Lin Yi Wang Sihao Wu Ying Sun Bowei Zhang huan Lv Xuemeng Ji Yang Lu Shuo Wang Tianjin Key Laboratory of Food Science and health School of MedicineNankai UniversityTianjin 300071China College of Food Science and Engineering Tianjin University of Science and TechnologyTianjin 300457China

Peanut allergy is considered to be a major health issue with global effects.To date,no effective curative approach has been applied for the therapy of the anaphylaxis resulting from the peanut allergens.The accurate and effective detection methods for the surveillance of allergens in food are still the primary strategies to avoid allergic diseases.In this study,nanobodies(Nbs)derived from the heavy-Chain only Antibodies(hCAbs)were selected against the general peanut protein extract through the unbiased strategy to facilitate the development of the sandwich ELISA for the detection and surveillance of peanut allergen contamination.The target antigen of the selected Nb was identified as peanut allergen Ara h 3,and a cross-reaction was observed with the member of Gly 1 from the Ara h 3 family.The applicability of the self-paired Nb P43 on the establishment of the immuno-assay was verified.A sandwich ELISA against peanut allergen was developed,which reached a linear range of 0.2-10.6μg/mL,and a limit of detection of 53.13 ng/mL.

关键词： Peanut allergen Ara h 3 Nanobody Sandwich ELISA Gly 1