A method of layer-by-layer gold nanoparticle hybridization in a quartz crystal microbalanceDNA sensing system used to detect dengue virus

Abstract

Dengue virus (DENV) is nowadays the most important arthropod-spread virus affectinghumans existing in more than 100 countries worldwide. A rapid and sensitive detectionmethod for the early diagnosis of infectious dengue virus urgently needs to bedeveloped. In the present study, a circulating-flow quartz crystal microbalance (QCM)biosensing method combining oligonucleotide-functionalized gold nanoparticles(i.e. AuNP probes) used to detect DENV has been established. In the DNA–QCMmethod, two kinds of specific AuNP probes were linked by the target sequences ontothe QCM chip to amplify the detection signal, i.e. oscillatory frequency change (ΔF) of the QCM sensor. The target sequences amplified from the DENV genome actas a bridge for the layer-by-layer AuNP probes’ hybridization in the method.Besides being amplifiers of the detection signal, the specific AuNP probes used inthe DNA–QCM method also play the role of verifiers to specifically recognizetheir target sequences in the detection. The effect of four AuNP sizes on thelayer-by-layer hybridization has been evaluated and it is found that 13 nm AuNPscollocated with 13 nm AuNPs showed the best hybridization efficiency. Accordingto the nanoparticle application, the DNA–QCM biosensing method was able todetect dengue viral RNA in virus-contaminated serum as plaque titers being 2 PFU ml−1 and a linearcorrelation (R2 = 0.987) ofΔF versus virustitration from 2 × 100 to 2 × 106 PFU ml−1 was found. The sensitivity and specificity of the present DNA–QCM method withnanoparticle technology showed it to be comparable to the fluorescent real-time PCRmethods. Moreover, the method described herein was shown to not require expensiveequipment, was label-free and highly sensitive.