Abdullin T.

Saifullina D.

Shakhmaeva I.

Materials Science and Engineering C

We developed metabolite-sensitive electrochemical sensors on the basis of electrodes modified with a thick film of carbon nanotubes. Modified electrodes provide efficient pre-adsorption of cellular metabolites and their sensitive detection using anodic square-wave voltammetry. On the electrode surface both adhered and non-adhered human cells produce three oxidation peaks at the potentials of + 0.82, + 1.05, and + 1.17 V attributed to three groups of cellular metabolites: amino acid-derived antioxidants including glutathione, guanine nucleotides, and also adenine nucleotides including ATP. The electrochemical response was well correlated with cell viability, intracellular ATP level and induction of apoptosis, as determined by independent assays. Developed sensors allow for robust and cost-effective assessment of ATP in cells in contrast to enzyme-based electrodes and conventional bioluminescent assay. Results can be used for rapid analysis of human cells for the purpose of medical diagnostics, transplantology, and toxicological screening. Additionally, we combined modified electrodes with human cells entrapped in agarose matrix. The resulting biosensor allowed for electrochemical monitoring of metabolic activity and death of cells within polymeric matrix that is of interest for tissue engineering applications. © 2012 Elsevier B.V. All rights reserved

Kazan Federal University Digital Repository

Assessment of metabolic activity of human cells in solution and in polymer matrixwith the use of metabolite-sensitive sensorsDiana V. Saifullina, Irina I. Shakhmaeva, Timur I. Abdullin ⁎Kazan (Volga Region) Federal University, 18 Kremlyovskaya St., 420008 Kazan, Republic of Tatarstan, Russiaa b s t r a c ta r t i c l e i n f oArticle history:Received 22 August 2011Received in revised form 27 February 2012Accepted 2 May 2012Available online 9 May 2012Keywords:Human cellsMetabolitesAntioxidantsATPElectrochemical (bio-)sensorsApoptosisBiomaterialsWe developed metabolite-sensitive electrochemical sensors on the basis of electrodes modiﬁed with a thickﬁlm of carbon nanotubes. Modiﬁed electrodes provide efﬁcient pre-adsorption of cellular metabolites andtheir sensitive detection using anodic square-wave voltammetry. On the electrode surface both adheredand non-adhered human cells produce three oxidation peaks at the potentials of +0.82, +1.05, and+1.17 V attributed to three groups of cellular metabolites: amino acid-derived antioxidants including gluta-thione, guanine nucleotides, and also adenine nucleotides including ATP. The electrochemical response waswell correlated with cell viability, intracellular ATP level and induction of apoptosis, as determined by inde-pendent assays. Developed sensors allow for robust and cost-effective assessment of ATP in cells in contrastto enzyme-based electrodes and conventional bioluminescent assay. Results can be used for rapid analysis ofhuman cells for the purpose of medical diagnostics, transplantology, and toxicological screening. Additional-ly, we combined modiﬁed electrodes with human cells entrapped in agarose matrix. The resulting biosensorallowed for electrochemical monitoring of metabolic activity and death of cells within polymeric matrix thatis of interest for tissue engineering applications.© 2012 Elsevier B.V. All rights reserved.1. IntroductionBiochemical analysis of mammalian cells is carried out in variousbiomedical studies [1,2]. Developmental and pathological processesin living cells are accompanied with a change in composition of plas-ma membrane components and intracellular metabolites [3]. Study-ing biochemical parameters of living cells is of particular interest inmedical diagnostics of many human disorders [1] as well as for drugand toxicant screening [4].Fluorescent probes/labeled antibodies are routinely used for thedetection of individual components in ﬁxed and live cell sampleswith the aid of optical microscopy [5]. Since ﬂow cytometry hasbeen introduced for quantitative ﬂuorescent analysis it became an es-sential technique for identiﬁcation and assessment of isolated cells[6,7].Other modern techniques involve chromatographic separation ofcell metabolites followed by their high-precision determination bymeans of mass spectrometry. Combined chromatography/mass spec-trometry is an irreplaceable tool for metabolome research when in-formative analysis of metabolite proﬁles is required [1,8]. However,limitations of this tool, e.g. high cost, laboriousness, inapplicabilityto live cell assays, complicate its introduction to some biomedical ap-plications including routine clinical diagnostics.Cellular metabolites can be detected without separation by the useof spectroscopic techniques such as Raman spectroscopy. This tech-nique allows for direct assessment of biochemical composition of livingcells by Raman spectra under physiological conditions. Changes inRaman spectra provide valuable information about normal and patho-logical cellular processes [9,10]. However, Raman analysis of biologicalmatter requires qualiﬁed interpretation of acquired data involving rela-tively complex mathematical methods.During the last decade many efforts have been made to developmore handy tools for rapid assessment of key cellular metabolites. Apromising platform for solving this problem is electrochemical sen-sors, especially voltammetric ones, which are characterized by selec-tivity, time efﬁciency and low cost of an analysis [11]. Electrochemicalsensors can be used for direct detection of a wide range of biomole-cules which are oxidized or reduced on the surface of electrodes. Incertain cases the electrode is coupled with enzyme(s) to performhighly selective quantiﬁcation of a metabolite of interest in biologicalsample [12]. To date, different electrochemical assays have been pro-posed for detection of catecholamines and nitric oxide on chemicallymodiﬁed electrodes [13–15] and also glutamate with the use of gluta-mate oxidase based sensors [16,17].A reliable parameter of energetic status, viability and induction ofapoptosis of human cells is intracellular ATP level [18–20]. Whereassome existing studies show the possibility of electrochemical detec-tion of some unidentiﬁed metabolites in cells [21–23] they do notconsider ATP and other adenine nucleotides which were shown tobe oxidized with high overpotential at about +1.2 V [24]. ForMaterials Science and Engineering C 32 (2012) 1843–1848⁎ Corresponding author. Tel.: +7 843 2337821; fax: +7 843 2387121.E-mail address: tabdulli@gmail.com (T.I. Abdullin).0928-4931/$ – see front matter © 2012 Elsevier B.V. All rights reserved.doi:10.1016/j.msec.2012.05.001Contents lists available at SciVerse ScienceDirectMaterials Science and Engineering Cj ourna l homepage: www.e lsev ie r .com/ locate /msec

Assessment of metabolic activity of human cells in solution and in polymer matrix with the use of metabolite-sensitive sensors

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