禽流感病毒流式微球量子点探针免疫诊断新方法

采用微波法水相中合成羧基化的绿色量子点,通过羧基与禽流感单克隆抗体氨基的共价结合,制备了检测禽流感病毒的探针,并结合流式微球技术,建立了量子点生物探针流式微球免疫检测禽流感病毒的新方法.以聚苯乙烯微球为蛋白质载体,将多克隆抗体包被到荧光微球上,依次加入待测抗原和量子点生物探针,形成双抗体夹心复合物,用流式细胞仪进行检测.实验结果表明,多抗和单抗的最佳质量浓度分别为92和4 mg/L,检测禽流感病毒比双抗体夹心ELISA灵敏16倍,比FITC标记单抗检测方法灵敏4倍.对阳性尿囊液的检测与ELISA呈现良好的相关性,不与鸡传染性支气管炎病毒、鸡马力克氏病毒、新城疫病毒等发生交叉反应.

A Novel Immunoassay of Detecting Avian Influenza Virus by Cytometry Using Quantum Dots Conjugation Probe and Flowing Microsphere Carrier

An aqueous solution of colloidal CdTe quantum dots(QDs)was prepared in the presence of MPA as a stabilizer.And QDs was used as fluorescent molecular probe after conjugating with a monoclonal antibody of avian influenza virus(AIV).An immunoassay was developed for determination of AIV by cytometry with flowing fluorescence-coded microsphere as protein carrier.Polyclonal antibody(PcAb) was coupling with microsphere,then antigen and DQs labeled McAb(McAb-Qds)were added sequencely to form the sandwich of double antibodies,and the detection was performed by flow cytometry.PcAb concentration of 92 mg/L and McAb concentration of 4 mg/L were selected as the optimization working concentrations.Under the optimal conditions,the results indicated that,comparing with traditional methods,the proposed method was 16 times more sensitive than ELISA and 4 times than FITC conjugated method.The method showed well relative with the ELISA method,and no cross-reaction occured with virus(NDV).