Department of Chemistry, Alfaisal University, Al Zahrawi Street, Al Maather, Al Takhassusi Road, Riyadh 11533, Saudi Arabia.
Department of Chemistry, Alfaisal University, Al Zahrawi Street, Al Maather, Al Takhassusi Road, Riyadh 11533, Saudi Arabia; King Faisal Specialist Hospital and Research Center, Zahrawi Street, Al Maather, Riyadh 12713, Saudi Arabia.
Biosens Bioelectron. 2017 May 15;91:169-174. doi: 10.1016/j.bios.2016.12.020. Epub 2016 Dec 10.
Food allergy has increased rapidly in recent years affecting millions of people worldwide. With the increased consumption of packed food nowadays, a sensitive, accurate and rapid screening method for potential food allergens has become an urgent need in order to protect the sensitive consumers from life-threatening reactions. The current detection methods for food allergens are mostly based on immunoassays which are costly and times-consuming. In this work, we developed an aptamer/graphene-based electrochemical biosensor for on-step, sensitive and low cost detection of β-lactoglobulin (β-LG) milk protein, one of the most common food allergens specially in infants. The selection of DNA aptamers against the two β-LG variants A and B was successfully realised using systemic evolution of ligands by exponential enrichment (SELEX) method. Among the selected aptamers, BLG14 aptamer sequence has shown high affinity and specificity to both β-LG A and B with dissociation constants (Ks) of 82 and 80nM, respectively as calculated using fluorescence binding assays. The aptamer was then integrated in a voltammetric biosensor utilizing graphene-modified screen printed carbon electrodes. The binding is monitored by following the change in the square wave voltammetry (SWV) reduction peak signal of ferrocyanide/ferricyanide redox couple due to the removal of the negatively charged aptamers from the surface upon protein binding. This one-step "signal on" biosensor takes 20min for the sensitive and selective detection of β-LG without using any labelling or sophisticated designs. The method was also tested in spiked food sample extract achieving good recovery percentage. The integration of the novel aptamer in the graphene biosensor allows a promising way for cost-effective, rapid and sensitive on-site detection of milk allergen in food stuff.
近年来，食物过敏的发病率迅速上升，影响了全球数百万人。由于如今人们对包装食品的消费增加，因此需要一种灵敏、准确和快速的筛选方法来检测潜在的食物过敏原，以保护敏感消费者免受危及生命的反应。目前用于检测食物过敏原的方法大多基于免疫测定法，这种方法既昂贵又费时。在这项工作中，我们开发了一种适体/基于石墨烯的电化学生物传感器，用于一步、灵敏和低成本检测β-乳球蛋白（β-LG）牛奶蛋白，β-LG 是最常见的食物过敏原之一，尤其是在婴儿中。使用指数富集的配体系统进化（SELEX）方法成功地选择了针对两种β-LG 变体 A 和 B 的 DNA 适体。在所选的适体中，BLG14 适体序列对β-LG A 和 B 均显示出高亲和力和特异性，荧光结合测定法计算出的解离常数（Ks）分别为 82 和 80nM。然后，该适体被整合到利用石墨烯修饰的丝网印刷碳电极的伏安生物传感器中。通过跟踪亚铁氰化物/铁氰化物氧化还原对的方波伏安（SWV）还原峰信号的变化来监测结合，由于结合蛋白质后，带负电荷的适体从表面上被去除，因此该信号发生变化。这种一步“信号开启”生物传感器在不使用任何标记或复杂设计的情况下，20 分钟即可灵敏且选择性地检测β-LG。该方法还在添加食物样品提取物中进行了测试，实现了良好的回收率。将新型适体集成到石墨烯生物传感器中为在食品中经济高效、快速且灵敏地现场检测牛奶过敏原提供了一种有前途的方法。
