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Fluorescent aptamer-based assay for thrombin with large signal amplification using peroxidase mimetics

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Abstract

This article describes an aptamer-based thrombin assay using a hemin-based peroxidase mimetic for signal amplification. Thrombin is recognized by an immobilized primary aptamer (G1-quadruplex). The G1-quadruplex/hemin complex formed quenches the fluorescence of CdTe quantum dots (QDs) due to photoinduced electron transfer (PET). In the next step, thrombin is associated with a secondary aptamer (G2-quadruplex) consisting of Pt nanoparticles (NPs), G2-quadruplex and hemin to form a sandwich structure. Both the G1-quadruplex/hemin complex and the Pt NPs/G2-quadruplex/hemin complex associated with thrombin act as enzyme mimetics and catalyze the oxidation of hydroquinone by H2O2 to form 2-hydroxy-p-benzoquinone (HPB). The HPB produced quenches the fluorescence of the CdTe QDs via a PET and an inner filter effect, thus causing large signal amplification. The effects were exploited to design a highly sensitive and selective thrombin assay. Under optimized conditions, a linear fluorescence response is achieved in the 0.05 pmol·L−1 to 10 nmol·L−1 concentration range, with a lower detection limit of 15 fmol·L−1. This approach, in our perception, represents a promising platform for sensitive detection of biomolecules for which appropriate aptamers can be found.

A sensitive fluorescent method was developed for the specific detection of thrombin. It is based on the photoinduced electron transfer (PET) induced fluorescence quenching of CdTe quantum dots (QDs) using enzyme mimetics for signal amplification.

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Acknowledgments

This work was supported by the National Natural Science Foundation of China (No.21275065), the Fundamental Research Funds for the Central Universities (JUSRP51314B) and the State Key Laboratory of Analytical Chemistry for Life Science of Nanjing University (KLACLS1008).

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Authors and Affiliations

  1. The Key Laboratory of Food Colloids and Biotechnology, Ministry of Education, School of Chemical and Material Engineering, Jiangnan University, Wuxi, 214122, People’s Republic China

    Guang-Li Wang, Xue-Lian Hu, Xiu-Ming Wu, Yu-Ming Dong & Zai-Jun Li

  2. State Key Laboratory of Analytical Chemistry for Life Science, Nanjing University, Nanjing, 210093, People’s Republic China

    Guang-Li Wang

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  1. Guang-Li Wang
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  2. Xue-Lian Hu
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Correspondence to Guang-Li Wang.

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Wang, GL., Hu, XL., Wu, XM. et al. Fluorescent aptamer-based assay for thrombin with large signal amplification using peroxidase mimetics. Microchim Acta 183, 765–771 (2016). https://doi.org/10.1007/s00604-015-1703-5

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  • DOI: https://doi.org/10.1007/s00604-015-1703-5

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