DNAzyme assisted recycling amplification method for ultrasensitive amperometric determination of lead(II) based on the use of a hairpin assembly on a composite prepared from nitrogen doped graphene, perylenetetracarboxylic anhydride, thionine and gold nanoparticles

A Pb(II)-DNAzyme is used in an amperometric method for the determination of Pb(II). The method is based on two feedback processes. In the first, the Pb(II)-DNAzyme initiates a reaction in presence of Pb(II) in a micro-tube to release a linear DNA (S₁). In the second, the S₁ triggers the recycling amplification between two types of hairpin-shaped DNA templates (H₁ and H₂) which consist of a primer sequence and a Pb(II)-DNAzyme substrate sequence. The Pb(II)-DNAzyme has excellent cleavage specificity toward the substrate sequence in S₁ that combined firstly with H₁ and then is linked to H₂. This process will connect H₁ and H₂. After hybridization with H₁ and H₂ to form two DNA complexes, S₁ is released and initiates the next recycling process. This results in efficient amplification. A glassy carbon electrode (GCE) was immersed into solution of HAuCl₄ to electrodeposit a layer of gold nanoparticles. This is followed by the assembly of the hairpin probe H₁ on the GCE. In addition, a nanohybrid consisting of 3, 4, 9, 10-Perylenetetracarboxylic acid (PTCA) and nitrogen-doped graphene (NG) was loaded with electroactive thionine (Thi) and gold to form nanoparticles of type NG-PTCA-Thi-Au. This is responsible for generating the amperometric signal (best measured at around -0.30 V vs. SCE) and also acts as the reducing agent for synthesizing the NG-PTCA-Thi-Au nanohybrid. H₂ is immobilized on NG-PTCA-Thi-Au to form a new tracer label. The concentration of Pb(II) in a solution can be quantified by determination of the amount of cleaved S₁. The method has high sensitivity and selectivity for Pb(II). The detection limit is 0.42 pM (S/N = 3), and the detection range extends from 1 pM to 1000 nM. Graphical abstract Schematic representation of electrodes for the determination of the lead ions (Pb²⁺). The sensor is using Pb²⁺-DNAzyme assisted recycling amplification based on hairpin assembly on a composite prepared from nitrogen doped graphene, perylenetetracarboxylic anhydride, thionine and gold nanoparticles (NG-PTCA-Thi-Au). This versatile platform expands studies on the detection of heavy metal ions.