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Aptamer-antibody sandwich assay for cytochrome c employing an MWCNT platform and electrochemical impedance

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Abstract

We report on a sensitive aptamer-antibody interaction-based assay for cytochrome c (Cyt c) using electrochemical impedance. 4-Amino benzoic acid is used for the oriented immobilization of aminated aptamers onto multi-walled carbon nanotubes on the surface of a screen-printed electrode via electrochemical grafting. Impedance was measured in a solution containing the redox system ferro/ferricyanide. The change in interfacial charge transfer resistance (Rct) experienced by the redox marker was recorded to confirm the formation of a complex between aptamer and the target (Cyt c). A biotinylated antibody against cytochrome c was then used in a sandwich type of assay. The addition of streptavidin conjugated to gold nanoparticles and signal enhancement by treatment with silver led to a further increase in Rct. Under optimized conditions, a detection limit as low as 12 pM was obtained. Cross-reactivity against other serum proteins including fibrinogen, BSA and immunoglobulin G demonstrated improved selectivity.

Sensitive and selective assay for cytochrome c protein using aptamer linked to multi-walled carbon nanotube screen printed electrode via diazonium electrochemical grafting and specific biotinylated antibody to improve selectivity. Detection can be based on electrochemical impedance spectroscopy, or using a streptavidin-gold nanoparticle conjugate.

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Acknowledgments

This research was partly supported by the Research Executive Agency (REA) of the European Union under Grant Agreement number PITN-GA-2010-264772 (ITN CHEBANA), by the Ministry of Science and Innovation (MCINN, Madrid, Spain) through the project CTQ2013-41577-P and by the Catalonia program ICREA Academia. Cristina Ocaña thanks the support of Ministry of Science and Innovation (MICINN, Madrid, Spain) for the predoctoral grant.

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Authors and Affiliations

  1. Sensors and Biosensors Group, Department of Chemistry, Universitat Autònoma de Barcelona, Edifici Cn, 08193 Bellaterra, Barcelona, Spain

    Cristina Ocaña & Manel del Valle

  2. Institute of Analytical Chemistry, Chemo- and Biosensors, University of Regensburg, 93053, Regensburg, Germany

    Sonja Lukic

Authors
  1. Cristina Ocaña
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  2. Sonja Lukic
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  3. Manel del Valle
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Correspondence to Manel del Valle.

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Ocaña, C., Lukic, S. & del Valle, M. Aptamer-antibody sandwich assay for cytochrome c employing an MWCNT platform and electrochemical impedance. Microchim Acta 182, 2045–2053 (2015). https://doi.org/10.1007/s00604-015-1540-6

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  • DOI: https://doi.org/10.1007/s00604-015-1540-6

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