DNA hybridization biosensors using polylysine modified SPCEs

Abstract

Two electrochemical DNA hybridization biosensors (genosensors) for the detection of a 30-mer sequence unique to severe acute respiratory syndrome (SARS) virus are described in this work. Both genosensors rely on the hybridization of the oligonucleotide target with its complementary probe, which is immobilized on positively charged polylysine modified screen-printed carbon electrodes (SPCEs), through electrostatic interactions. In one design, a biotinylated target is used and the detection of the hybridization reaction is monitored using alkaline phosphatase labeled streptavidin (S-AP). This enzyme catalyzes the hydrolysis of the substrate 3-indoxyl phosphate (3-IP) to indigo, which is then solubilized to indigo carmine and detected by means of cyclic voltammetry (CV). In the other design, the target is labeled using an Au(I) complex, sodium aurothiomalate, and the duplex formation is detected by measuring, for first time, the current generated by the hydrogen evolution catalyzed by the gold label. Using 30 min of hybridization time, a detection limit of 8 pM is calculated for the enzymatic genosensor. Although this good sensitivity cannot be reached with the metal label (0.5 nM), the use of this label allows a considerable decrease of the analysis time. Both genosensors do not require the modification of the oligonucleotide probe and using stringent experimental conditions (60 min of hybridization time and 50% formamide in the hybridization buffer) can discriminate between a complementary oligonucleotide and an oligonucleotide with a three-base mismatch.

Poly‐L‐lysine Coated Surfaces for Ultrasensitive Nucleic Acid Detection

“…Hybridization event has been monitored with methylene blue. The detection limit has been found as 1.69×10 −13 M. A genesensor to detect severe acute respiratory syndrome (SARS) virus has been prepared on poly‐L‐ysine modified screen‐printed carbon electrodes (SPCEs) . Enzyme catalyzed hydrolysis of the 3‐indoxyl phosphate (3‐IP) to indigo has been monitored by cyclic voltammetry.…”

“…In the first part of this study, we aim to prepare poly‐L‐lysine coated disposable pencil graphite electrodes (PLL/PGEs) and investigate the electrochemical behaviors of these PLL/PGEs. PLL coated surfaces have attracted great attention due to their positively charged amino groups (−NH 2 ) and negatively charged carboxyl groups (−COOH), versatility and ease of electrochemical polymerization . In addition, Poly‐L‐lysine/graphene (PLL/GN) and poly‐L‐lysine/multi‐walled carbon nanotubes (PLL/MWCNTs) nanocomposites coated PGEs were prepared in one‐pot synthesis and introduced for the first time in the literature.…”

Gold nanoparticle-based electrochemical magnetoimmunosensor for rapid detection of anti-hepatitis B virus antibodies in human serum

“…During the application of this potential, some gold atoms in the outer layers of AuNPs surface are transformed into Au (III) ions. These ions could also exert a catalytic effect on hydrogen evolution [Díaz-González et al, 2008] together with the significant number of AuNPs still remaining after the oxidation step. After that, the catalytic current generated by the reduction of the hydrogen ions is chronoamperometrically recorded and related to the quantity of the ␣-HBsAg IgG antibodies.…”

Influence of Graphene Oxide Concentration when Fabricating an Electrochemical Biosensor for DNA Detection

“…In the latest years, several pretreatment procedures for the SPCE surface aiming to improve the sensitivity and reproducibility of the results have been reported [43][44][45][46][47][48][49][50]. Improvement of the electrochemical performance of screen-printed carbon electrodes by UV/ozone modification was very recently described [43].…”