用于血清胰岛素测定的荧光共振能量传递(FRET)免疫检测法的建立

Development of a fluorescence resonance energy transfer(FRET)immunoassay for the measurement of serum insulin

目的研究探讨新型荧光染料Dylight(DL)和金纳米颗粒(AuNP)间荧光共振能量转移(FRET)效应,并将其与免疫竞争法结合构建一种新型胰岛素荧光免疫检测方法。方法分别将胰岛素抗原(Ag)和胰岛素抗体(Ab)与DL和AuNP进行偶联,形成DL-Ag偶联物和AuNP-Ab偶联物。利用它们间产生的FRET效应以及免疫竞争反应,建立针对胰岛素的新型荧光免疫检测方法。然后对该检测方法的检测性能进行评估和实际样品的应用研究。结果该荧光免疫检测方法不仅表现出较高的检测灵敏度(0.015 ng/mL),较快的检测时间(4 min)以及较好的检测特异性,而且成功地用于血清胰岛素的检测,回收率在96.9%~121.1%之间。结论新型荧光染料DL和AuNP间FRET效应可应用于血清胰岛素的荧光免疫检测。

Objective To investigate the fluorescence resonance energy transfer(FRET)effect between dylight(DL)and AuNP(AuNP),and to construct a new fluorescence immunoassay for insulin in combination with the immunocompetition method.Methods Insulin antigen(Ag)and insulin antibody(Ab)were conjugated with DL and AuNP respectively to form DL-Ag conjugate and AUNp-AB conjugate.A novel fluorescence immunoassay for insulin was developed on the basis of FRET effect and the immune competition response between them.Then the performance of the method was evaluated and its application in actual samples was explored.Results The fluorescence immunoassay showed high sensitivity(0.015 ng/mL),short measurement time(4 min)and good specificity.It was successfully used in the measurement of serum insulin,and the recovery was between 96.9%and 121.1%.Conclusion FRET effect between AuNP and DL can be applied to develop a fluorescence immunoassay for the measurement of serum insulin.