ORIGINAL PAPER Selection of DNA aptamers against Mycobacterium tuberculosis Ag85A, and its application in a graphene oxide-based fluorometric assay Najmeh Ansari 1 & Kiarash Ghazvini 2 & Mohammad Ramezani 3 & Mahin Shahdordizadeh 4 & Rezvan Yazdian-Robati 4 & Khalil Abnous 5 & Seyed Mohammad Taghdisi 6 Received: 24 July 2017 /Accepted: 9 November 2017 /Published online: 5 December 2017 # Springer-Verlag GmbH Austria, part of Springer Nature 2017 Abstract The Mycobacterium Ag85 complex is the major secretory protein of M. tuberculosis. It is a potential marker for early diagnosis of tuberculosis (TB). The authors have identified specific aptamers for Ag85A (FbpA) via protein SELEX using magnetic beads. After twelve rounds of selection, two aptamers (Apt8 and Apt22) were chosen from different groups, and their binding constants were determined by flow cytometry. Apt22 (labeled with Atto 647N) binds to FbpAwith high affinity (K d = 63 nM) and specificity. A rapid, sensitive, and low-cost fluorescent assay was designed based on the use of Apt22 and graphene oxide, with a limit of detection of 1.5 nM and an analytical range from 5 to 200 nM of FbpA. Keywords SELEX . Serum . Fluorescent assay . Limit of detection . Secretory protein . Quenching Introduction Tuberculosis (TB), caused by Mycobacterium tuberculosis, is still the major global health infectious disease that is respon- sible for high morbidity, especially in developing countries. Thesurvey in 2014 showed 9 million people fell ill with TB and 1.3 million M. tuberculosis deaths were occurred [1]. Emergence of multi-drug resistant (MDR) strains and co- infections with human immunodeficiency virus (HIV) cause high prevalence of TB [2]. Early diagnosis of TB is important for efficiently controlling the epidemic disease. Conventional diagnostic systems of TB rely on the myco- bacterial culture and smear microscopy of acid fast bacilli (AFB) [3]. These approaches have major drawbacks. AFB staining sensitivity is variable, ranging from 20% to 80% [4]. Traditional culture detection, which is BGold standard^ of clinical TB diagnosis requires extended incubation times, 6–8 weeks [5]. Also assays such as DNA hybridization, fluo- rescent antibody test, and polymerase chain reaction (PCR) have been developed for detection of TB [6, 7]. However, these methods need costly equipment, well-established labo- ratory, and highly trained personnel. Consequently, it is Electronic supplementary material The online version of this article (https://doi.org/10.1007/s00604-017-2550-3) contains supplementary material, which is available to authorized users. * Khalil Abnous abnouskh@mums.ac.ir * Seyed Mohammad Taghdisi taghdisihm@mums.ac.ir 1 Department of Microbiology and Virology, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad 91778-99191, Iran 2 Antimicrobial Resistance Research Center, Buali Research Institute, Department of Microbiology and Virology, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad 91778-99191, Iran 3 Nanotechnology Research Center, Mashhad University of Medical Sciences, Mashhad 91778-99191, Iran 4 Department of Pharmaceutical Biotechnology, School of Pharmacy, Mashhad University of Medical Sciences, Mashhad 91778-99191, Iran 5 Pharmaceutical Research Center, Mashhad University of Medical Sciences, Mashhad 91778-99191, Iran 6 Targeted Drug Delivery Research Center, Mashhad University of Medical Sciences, Mashhad 91778-99191, Iran Microchimica Acta (2018) 185: 21 https://doi.org/10.1007/s00604-017-2550-3