在前期已获得抗金黄色葡萄球菌肠毒素B (Staphylococcal enterotoxin B, SEB)纳米抗体的基础上,建立一种用于检测食品中SEB的酶联免疫吸附法(enzyme-linked immunosorbent assay, ELISA),为食品及农产品质量监管提供技术支持。优化表达条件后,通过大肠杆菌原核表达获得抗SEB的纳米抗体B7;以纳米抗体B7作为捕获抗体,以噬菌体展示的抗SEB纳米抗体B6为检测识别元件,建立夹心ELISA方法;以牛奶、牛肉及西瓜汁为样本对其进行方法学评价。结果显示,纳米抗体B7在大肠杆菌中成功表达,表达量为 6.2 mg/L;该方法在16~1 024 ng/mL具有良好的线性关系(R2=0.990 4),最低检出限为(9.58±0.07)ng/mL;该方法与SEC有42.18%的交叉,与SEA及另外3株金黄色葡萄球菌无明显交叉;牛奶、牛肉和西瓜汁的加标回收率分别为87.26%~108.43%,79.36%~106.56% 和 83.81%~99.43%。该方法以纳米抗体作为识别元件,可应用于实际食品及农产品中的SEB高灵敏检测,具有广阔的应用前景。
In this study, a method for detection of Staphylococcal enterotoxin B (SEB) in foodstuffs through anti-SEB nanobody-mediated ELISA was developed using milk, beef and watermelon juice as samples. The method exhibited a desirable linear relationship (R2=0.9904) in a concentration range from 16 to 1 024 ng/mL, and the detection minimum was (9.58±0.07) ng/mL. The method exhibited 42.18% crossover with S. enterotoxin C while there were no significant crossovers with S. enterotoxin A and other three strains of Staphylococcus aureus. The recoveries of SEB in milk, beef and watermelon juice were 87.26%-108.43%, 79.36%-106.56% and 83.81%-99.43%, respectively. In conclusion, the ELISA used anti-SEB nanobody as a recognition element could be an alternative method for highly sensitive detection of SEB.
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