Electrochemical detection of individual single nucleotide polymorphisms using monobase-modified apoferritin-encapsulated nanoparticles

An electrochemical approach for detection of individual single nucleotide polymorphisms (SNPs) based on nucleobase-conjugated apoferritin probe loaded with metal phosphate nanoparticles is reported. Coupling of the nucleotide-modified nanoparticle probe to the mutant sites of duplex DNA was induced by DNA polymerase I (Klenow fragment) to preserve Watson–Crick base-pairing rules. After sequential liquid hybridization of biotinylated DNA probes with mutant DNA and complementary DNA, the resulting duplex DNA helixes were captured to the surface of magnetic beads through a well known and specific biotin-streptavidin affinity binding. For signaling each of eight possible Single-nucleotide polymorphisms (SNPs), Pb, Cu, Cd and Zn phosphate-loaded apoferritin nanoparticle probes were linked to adenosine (A), cytidine (C), guanosine (G), and thymidine (T) mononucleotides, respectively. Monobase-conjugated apoferritin probes were coupled to the mutant sites of the formed duplex DNA in the presence of DNA polymerase. Electrochemical stripping analyses of the metals loaded in apoferritin nanoparticle probes provide a means for detection and quantification of mutant DNA. Each mutation captures different nucleotide-conjugated apoferritin probe and provide a distinct four-potential voltammogram, whose peak potentials reflect the identity of the mismatch. The method is sensitive enough to accurately determine AG mutation, as the most thermodynamically stable mismatch to detect, in the range of 50–600 pM. The proposed protocol provides a simple, fast, cost-effective, accurate and sensitive method for detection of SNPs.

► Apoferritin-encapsulated metal phosphate nanoparticles as electrochemical marker for detection of SNPs.
► Streptavidin-coated magnetic beads as a substrate for capturing biotinylated DNA probes.
► Voltammetric stripping analysis of metallic phosphate nanoparticles on a mercury coated screen printed electrode.
► Detection of all eight possible SNPs in a single voltammetric run.
► Possession of high sensitivity in detection of low concentrations of SNPs.