In this study, disruption and lyophilization-rehydration of dopamine-loaded liposome and its application in electrochemical DNA biosensor was investigated. The liposomes containing soyphosphatidylcholine and cholesterol were prepared through thin-layer hydration. First, an investigation was carried out to find an appropriate lysing agent for disruption of prepared liposomes. Differential pulse voltammetry, as a high sensitive electrochemical technique, was used along with a multi-walled carbon nanotubes modified glassy carbon electrode for sensitive electrochemical detection of released dopamine from disrupted liposomes. Various lysing agents were investigated and finally, the disruption of liposomes using methanol was selected without any surfactant, because of its least fouling effect. Then, lyophilization of dopamine-loaded liposomes was carried out using sucrose as cryoprotectant. The electrochemical studies of lyophilized liposomes showed that the remained dopamine in sucrose-protected liposomes was higher than sucrose-free liposomes. Furthermore, sucrose has no interference in electrochemical studies. Then, with the addition of biotin-X-DHPE to liposome formulation, the lyophilized sucrose protected dopamine-loaded biotin-tagged liposomes were prepared and the feasibility of application of them in electrochemical DNA biosensor was investigated as signal enhancer and verified for detection of oligonucleotides.
In this study, development of an electrochemical DNA biosensor based on a hepatitis C virus probe was introduced using an electrochemically pre-treated pencil graphite electrode (PPGE) as a transducer and Nile blue (NB) as an electroactive indicator. At first, the electrochemical behaviour of NB on the PPGE was investigated. Then, elucidating the specific interaction of NB with each of the nucleotides of DNA, the interactions of NB with oligonucleotides containing only one base type were studied. In this study, four 18-mer oligonucleotides of poly A, poly T, poly C, and poly G were used as probe and were used as related complementary/noncomplementary sequences in the hybridization section. The extent of hybridization was evaluated based on the difference between DPV signals of NB accumulated on the probe-PPGE and NB accumulated on the probe-target-PPGE. Then, the developed biosensor was applied successively for the detection of short sequences of Hepatitis C 3a virus (14 pic). The hybridization between the probe (14 pic) and its complementary sequence (C 2 Comp) as the target was studied. Some hybridization experiments with noncomplementary oligonucleotides also showed that the suggested DNA sensor responds selectively to the target. Diagnostic performance of the developed biosensor was described and the detection limit was found to be 1.1 nM with a relative standard deviation of 3.5 % in the 0.1-M Tris-HCl buffer solution (pH 7.0) containing 20 mM NaCl.
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