聚合酶活性信号检测地高辛方法的建立

Establishment of a digoxin detection method by polymerase Activity signal

目的建立以聚合酶活性信号为检测对象的反应体系,使用具有高灵敏度和精密度的均相分析方法,实现地高辛血药浓度的自动化检测。方法设计特异的DNA模板及其引物,引物上特定位点连接有Digoxingenin-11-dUTP（Dxin）,使得样品中地高辛（Dig）与引物上的Dxin在抗Dig抗体存在时形成竞争反应,影响DNA聚合酶与引物结合,从而根据合成的双链量即可检测出样品Dig浓度。结果选择D2p作为Dxin在引物上的结合位点,17nmol/L模板及引物浓度、33nmol/L的抗Dig抗体浓度。试剂可报告范围为0.3～39.0ng/mL,CV〈10%。结论聚合酶活性信号检测技术可以实现高灵敏度的地高辛均相分析。

Objective To establish a reaction system with the polymerase activity signal as the detection object and to realize the automatic detection of digoxin（Dig）blood concentration by using the homogeneous phase assay method with high sensitivity and high precision.Methods The specific DNA template and its primer which conjugated with Digoxingenin-11-dUTP（Dxin）at specific site were designed.Dig in serum sample was formed the competing reaction with Dxin on the primer when anti-Digoxin antibody existed,which influenced the combination of DNA polymerase with primer,thus the dig content in the sample could be detected according to the synthesized double chain amount.Results D2 p as the binding site of Dxin on the primer,17nmol/L template,primer concentration and 33nmol/L anti-Dig concentration were selected.The reportable range of reagent was 0.3-39.0ng/mL,CV〈10%.Conclusion The polymerase signal detection technology can realize the Dig homogeneous phase analysis with high sensitivity.