title: Real-time electrochemical monitoring of adenosine triphosphate in the picomolar to micromolar range using graphene-modified electrodes.
abstract: We report on a competitive electrochemical detection system that is free of wash steps and enables the real-time monitoring of adenosine triphosphate (ATP) in a quantitative manner over a five-log concentration range. The system utilizes a recognition surface based on ATP aptamer (ATPA) capture probes prebound to electroactive flavin adenine dinucleotide (FAD) molecules, and a signaling surface utilizing graphene (Gr) and gold nanoparticle (AuNP) modified carbon paste electrode (Gr-AuNP-CPE) that is optimized to enhance electron-transfer kinetics and signal sensitivity. Binding of ATP to ATPA at the recognition surface causes the release of an equivalent concentration of FAD that can be quantitatively monitored in real time at the signaling surface, thereby enabling a wide linear working range (1.14 × 10(-10) to 3.0 × 10(-5) M), a low detection limit (2.01 × 10(-11) M using graphene and AuNP modified glassy carbon), and fast target binding kinetics (steady-state signal within 12 min at detection limit). Unlike assays based on capture probe-immobilized electrodes, this double-surface competitive assay offers the ability to speed up target binding kinetics by increasing the capture probe concentration, with no limitations due to intermolecular Coulombic interactions and nonspecific binding. We utilize the real-time monitoring capability to compute kinetic parameters for target binding and to make quantitative distinctions on degree of base-pair mismatch through monitoring target binding kinetics over a wide concentration range. On the basis of the simplicity of the assay chemistry and the quantitative detection of ATP within fruit and serum media, as demonstrated by comparison of ATP levels against those determined using a standard high-performance liquid chromatography (HPLC)-UV absorbance method, we envision a versatile detection platform for applications requiring real-time monitoring over a wide target concentration range.
authors.affiliation: Department of Electrical and Computer Engineering, University of Virginia, Charlottesville, Virginia 22904, USA.
authors.forename: Bankim J
authors.initials: BJ
authors.lastname: Sanghavi
authors.forename: Sarita
authors.initials: S
authors.lastname: Sitaula
authors.forename: Mark H
authors.initials: MH
authors.lastname: Griep
authors.forename: Shashi P
authors.initials: SP
authors.lastname: Karna
authors.forename: Mehnaaz F
authors.initials: MF
authors.lastname: Ali
authors.forename: Nathan S
authors.initials: NS
authors.lastname: Swami
mesh_terms: D000255:Adenosine Triphosphate / Q000032:analysis / Q000737:chemistry*; D003199:Computer Systems; D055664:Electrochemical Techniques / Q000379:methods*; D004566:Electrodes; D006108:Graphite / Q000737:chemistry*; D006801:Humans; D008297:Male
chemical_list: D006108:Graphite; D000255:Adenosine Triphosphate