An electrochemical detection method for ATP based on homogeneous biorecognition reaction and TdT-induced polymerization signal amplification
Energy levels in organisms are closely linked to metabolic diseases, with ATP—the universal energy currency—playing a central role in this relationship. This paper presents a novel electrochemical biosensor for ATP, integrating homogeneous bio-recognition reactions with terminal deoxynucleotidyl transferase (TdT)-mediated polymerization to enable precise ATP detection in metabolic research. Initially, a DNA oligonucleotide composed of a camouflaged aptamer and exposed hairpin DNA was designed to react homogeneously with ATP in the presence of exonuclease I (Exo I). This reaction generates blunt-ended hairpin DNA, which can activate the assembly of an ordered DNA-rich nanostructure layer on the gold electrode. Subsequently, the DNA strand is elongated through TdT-induced polymerization in the presence of deoxynucleotide triphosphates (dNTPs). A significant amount of Ru(NH 3 ) 6 3+ can be adsorbed onto the negatively charged DNA phosphate backbone via strong electrostatic interactions, effectively amplifying the detection signal and enhancing the sensor's performance. Under optimal conditions, the system exhibits excellent linearity over the target concentration range of 10 −2 to 10 3 nM, with a detection limit as low as 8.37 pM, enabling the detection of trace amounts of ATP. The biosensor demonstrates high stability and reliability, with recovery rates ranging from 97.8 % to 110 %. This method serves as a diagnostic tool to assess organismal metabolic status by monitoring ATP levels, enabling the tracking of metabolic disease progression. By integrating molecular monitoring with clinical diagnosis, it offers a potential approach for the preventing and intervention of metabolic diseases.
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