基于荧光染料和氧化石墨烯同步荧光法同时检测多种流感病毒亚型

Synchronous Fluorescence Detection of Three Influenza Virus Subtypes Based on the Fluorescent Dyes and Graphene Oxide

分别采用三种不同发射波长的荧光染料,通过共价偶联的方式分别标记三种流感病毒亚型H1N1、H5N1和H9N2的抗体,再利用氧化石墨烯猝灭所标记染料的荧光。当将荧光标记抗体和氧化石墨烯一并加入到流感病毒溶液中时,由于抗原和抗体之间的特异性相互作用,病毒会和抗体作用而使得氧化石墨烯远离荧光染料,染料的荧光得以恢复。通过恢复的荧光发射波长位置和荧光强度,可以定性和定量检测三种不同的流感病毒亚型。在最佳实验条件下,对三种流感病毒亚型H1N1、H5N1和H9N2进行同时检测,H1N1的检出限为0.48ng/mL,线性范围为1～18ng/mL;H5N1的检出限为0.46ng/mL,线性范围为1-18.5ng/mL;H9N2的检出限为0.42ng/mL,线性范围为1～16ng/mL。该方法具有较好的稳定性、重现性和灵敏度,可实现多个流感病毒亚型的分型和同时检测。

Three types of influenza virus subtype H1N1, H5N1, H9N2 antibodies were covalently labeled by three fluorescent tags of different emission wavelength. Graphene oxide （GO） was used as a fluorescence quencher towards to the labeled dyes. By adding these fluorescent labeled antibodies and GO into the flu virus solution, due to the specificity of the interaction between antigen and antibody, viruses and antibody interaction will induce GO to get away from the fluorescent dye, which leads to the fluorescence intensity restored. By the restoring fluorescence intensity, it can realize the qualitative and quantitative detection of three influenza virus subtypes. Under the optimum experimental conditions, the three kinds of influenza virus subtypes were detected. The detection limit is 0.48 ng/mL with the linear range of 1 to 18 ng/mL for HIN1 virus,0.46 ng/mL with the linear range of 1-18.5 ng/mL for H5N1 virus,and 0.42 ng/mL,with the linear range of 1 to 16 ng/mL for H9N2 virus,respectively. This method has good stability, sensitivity and reproducibility, and can realize multiple virus subtypes classification and detection at the same time.