量子点标记荧光免疫法检测过氧化氢酶

Immunofluorescence by Using Quantum Dots Label for Detecting of Catalase

通过制备量子点标记过氧化氢酶的荧光探针,建立过氧化氢酶荧光免疫分析方法。通过碳二亚胺法,合成了过氧化氢酶-量子点荧光探针,对活化剂添加量、体系pH等荧光探针的合成条件及免疫分析检测中抗体浓度、封闭液BSA浓度、反应时间等参数进行优化,用灵敏度、批内变异、批间变异和加标回收率指标对方法进行评价。结果表明,活化剂的最佳添加量10μL、体系最佳pH 7.4、抗体的最佳浓度1μg/mL、封闭液BSA最佳浓度1%、反应时间60 min。方法检测范围为1~1000μg/mL,相关系数R2为0.9942,最低检测限为2.5×10-2μg/mL,批内、批间变异系数低于15%,具有良好特异性,样品的加标回收率高于90%。方法操作简单、准确度高、线性范围宽,为过氧化氢酶含量检测提供方向。

A method for fluorescence immunoassay of catalase was established by preparing a fluorescent probes with quantum dot labeled catalase.The catalase-quantum dot fluorescent probe was synthesized by the carbodiimide method,and the parameters were optimized about the synthesis conditions of fluorescent probe and immunoassay condition,including the addition amount of activator,system pH,antibody concentration,concentration of blocking solution BSA,reaction time and so on.The method was evaluated by sensitivity,intra-assay variation,inter-assay variation and standard recovery experiment.The results showed the optimal addition amount of activator 10μL,the optimal pH of the system 7.4,the optimal concentration of antibody 1μg/mL,the optimal concentration of BSA in blocking solution 1%,and the reaction time 60 min.The detection range of this method was 1-1000μg/mL;The correlation coefficient R2 was 0.9942;The detection limits was 2.5×10-2μg/m L;The intra-assay and inter-assay coefficient of variation was less than 15%;Good specificity was made;The standard recovery of samples was more than 90%.The method was simple in operation,high in accuracy and wide in linear range,and a new ideal for the detection of catalase content was provided.