RESEARCH ARTICLE A fluorescent aptasensor for analysis of adenosine triphosphate based on aptamer–magnetic nanoparticles and its single‐ stranded complementary DNA labeled carbon dots Zeinab Saberi | Behzad Rezaei | Taghi Khayamian Department of Chemistry, Isfahan University of Technology, Isfahan, Islamic Republic of Iran Correspondence Behzad Rezaei, Department of Chemistry, Isfahan University of Technology, Isfahan, 84156–83111 Islamic Republic of Iran. Email: rezaei@cc.iut.ac.ir Abstract A new fluorimetric aptasensor was designed for the determination of adenosine triphosphate (ATP) based on magnetic nanoparticles (MNPs) and carbon dots (CDs). In this analytical strategy, an ATP aptamer was conjugated on MNPs and a complementary strand of the aptamer (CS) was labeled with CDs. The aptamer and its CS were hybridized to form a double helical structure. The hybridized aptamers could be used for the specific recognition of ATP in a biological complex matrix using a strong magnetic field to remove the interfering effect. In the absence of ATP, no CDs–CS could be released into the solution and this resulted in a weak fluorescence signal. In the presence of ATP, the target binds to its aptamer and causes the dissociation of the double helical structure and liberation of the CS, such that a strong fluorescence signal was generated. The increased fluorescence signal was proportional to ATP concentration. The limit of detection was estimated to be 1.0 pmol L –1 with a dynamic range of 3.0 pmol L –1 to 5.0 nmol L –1 . The specific aptasensor was applied to detect ATP in human serum samples with satisfactory results. Moreover, molecular dynamic simulation (MDS) studies were used to analyze interactions of the ATP molecule with the aptamer. KEYWORDS adenosine triphosphate, aptamer, carbon dots, complementary strand of aptamer 1 | INTRODUCTION Pre‐treatment steps are very important in removing matrix effects when determining trace amounts of analytes in environmental and biological samples before measurement and analysis. As magnetic nanoparticles (MNPs) show superparamagnetic characteristics, they can be easily recovered and separated from a sample matrix with an external magnetic field, thus reducing possible interference and improving the figures of merit. In recent years, modified MNPs attached to aptamers as sorbents have been an interesting area of study for the analytical community. A high surface area to volume ratio, the advantage of MNPs, improves extraction efficiency and provides a simple and rapid extraction method without the use of filters. [1] Single‐stranded nucleic acid (ssDNA or ssRNA) molecules called aptamers are able to bind pre‐selected targets including proteins, peptides, small organic compounds and ions with high affinity and specificity based on their three‐dimensional (3D) structures. Several advantages of aptamers, such as intrinsic selectivity, affinity, simplicity, chemical stability and less susceptibility to denaturation, make them very good options for biosensors in the biomedical and biotechnology research fields. [2–4] In addition, the use of aptamer‐conjugated MNPs has been investigated previously. [5,6] Adenosine triphosphate (ATP) is a nucleoside triphosphate, and is known as a major chemical signaling agent. It plays a vital role in energy metabolism and signal transduction. ATP is often known as the molec- ular unit of currency of intracellular energy transfer. Therefore, ATP levels can be utilized to assess cell viability, injury, proliferation, and inhibition induced by various biological agents or small molecule drugs. [7] To date, different analytical methods have been reported for ATP analysis, including electrochemical assays [8,9] , chromatogra- phy [10] , electrochemiluminescence assays [11,12] , or fluorescence Abbreviations used: APTES, 3‐aminopropyl)triethoxysilane; CS, complementary strand; DLS, dynamic light scattering; EDC, 1‐ethyl‐3‐(3‐dimethyllaminopropyl) carbodiimide hydrochloride; LGA, Lamarckian genetic algorithm; LOD, limit of detection; MDS, molecular dynamic simulation; MNP, magnetic nanoparticle; NHS, N‐hydroxysulfosuccinimide sodium salt; PBS, phosphate buffer solution; PDB, program database; TEM, transmission electron micrograph. Received: 26 August 2017 Revised: 16 October 2017 Accepted: 24 December 2017 DOI: 10.1002/bio.3457 Luminescence. 2018;1–7. Copyright © 2018 John Wiley & Sons, Ltd. wileyonlinelibrary.com/journal/bio 1