A single-step method for direct detection of genomic DNA targets is demonstrated. The method is based on the use of thiolated linear single-stranded DNA (ssDNA), of respectively 20 and 21 nts, anchored onto gold surfaces. Different concentrations of ssDNA solutions, ranging from 38.8µM to 0.08µM, were employed, obtaining surface probe density between 1.0 x 1013 and 2.2 x 1012 molecules/cm2. The kinetics of ssDNA immobilization and the subsequent HBV-clone recognition process were studied by means of a Quartz Crystal Microbalance with Dissipation Monitoring (QCM-D) technique and Atomic Force Microscopy. The surface density of the thiolated ss DNA probes was found to affect strongly the efficiency of Hepatitis B Virus (HBV) clone detection and recognition. A critical surface density has been found to occur at about 4.0 x 1012 molecules/cm2 of oligonucleotide probes to obtain an optimal recognition process, giving rise to a specific and fast HBV-clone recognition, ensuring picomolar (pM) sensitivity without any DNA amplification steps.

Single-Step Label Free Hepatitis B Virus Detection by Piezoelectric Biosensor

MARLETTA, Giovanni
2015

Abstract

A single-step method for direct detection of genomic DNA targets is demonstrated. The method is based on the use of thiolated linear single-stranded DNA (ssDNA), of respectively 20 and 21 nts, anchored onto gold surfaces. Different concentrations of ssDNA solutions, ranging from 38.8µM to 0.08µM, were employed, obtaining surface probe density between 1.0 x 1013 and 2.2 x 1012 molecules/cm2. The kinetics of ssDNA immobilization and the subsequent HBV-clone recognition process were studied by means of a Quartz Crystal Microbalance with Dissipation Monitoring (QCM-D) technique and Atomic Force Microscopy. The surface density of the thiolated ss DNA probes was found to affect strongly the efficiency of Hepatitis B Virus (HBV) clone detection and recognition. A critical surface density has been found to occur at about 4.0 x 1012 molecules/cm2 of oligonucleotide probes to obtain an optimal recognition process, giving rise to a specific and fast HBV-clone recognition, ensuring picomolar (pM) sensitivity without any DNA amplification steps.
thiolated single stranded DNA; probe surface density; quartz crystal microbalance
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/20.500.11769/18628
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