Development of monoclonal antibodies against α-lactalbumin and establishment of qualitative detection for α- lactalbumin with dot blotting assay
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摘要: 以α-乳白蛋白为抗原免疫Balb/c小鼠,采用淋巴细胞杂交瘤技术制备抗α-乳白蛋白单克隆抗体;采用碳二亚胺法,并通过工艺优化,将单抗与量子点高效偶联;在此基础上,建立了基于斑点印迹技术定性检测乳制品中过敏原α-乳白蛋白的方法。结果表明:采用杂交瘤技术制备的单克隆抗体特异性好,与牛奶中其他蛋白无交叉反应;当量子点、1-（3-二甲氨基丙基）-3-乙基碳二亚胺盐酸盐（EDC）、N-羟基丁二酰亚胺（NHS）、单抗摩尔比为1∶10∶6∶4时,量子点与单抗有较好的偶联效果;以量子点标记单抗建立的斑点印迹方法可直观、快速的定性分析乳制品中过敏原α-乳白蛋白。因此,该方法具有一定的实用价值。Abstract: The monoclonal antibodies (mAbs) against bovine α-lactalbumin (α-la) were prepared by immunning Balb/c mice and the lymphocyte hybridoma technique. The mAbs were covalently conjugated with the CdSe/ZnS quantum dots (QDs) by the carbodiimide crosslinking method, and then the coupling conditions were optimized. Furthmore, a dot blotting method based on QD-mAb conjugates was established to qualitatively detect α-La in different commercial dairy products. The results showed that the prepared mAbs had high specificity and no cross-reaction. A good coupling effect was confirmed when the molar ratio of QD, N- (3-Dimethylaminopropyl) -N'-ethylcarbodiimide hydrochloride (EDC) , N-Hydroxysuccinimide (NHS) , monoclonal antibody was 1∶10∶6∶4. The developed method based on QD-mAb conjugates could be used to visual, rapid qualitative analyze α-La in dairy products. Accordingly, the method possesses certain practical value.
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Keywords:
- α-lactalbumin /
- monoclonal antibody /
- quantum dots /
- dot blotting /
- qualitative detection
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