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Affinity binding-mediated fluorometric protein assay based on the use of target-triggered DNA assembling probes and aptamers labelled with upconversion nanoparticles: application to the determination of platelet derived growth factor-BB

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Abstract

The target-triggered DNA assembling probe is presented for highly selective protein detection. Target-triggered DNA assembling is used in an amplification strategy based on affinity binding for identification and determination of proteins in general. Specifically, it was applied to the platelet derived growth factor-BB (PDGF-BB). A hairpin DNA (H-DNA) probe was designed containing (a) an aptamer domain for protein recognition and (b) a blocked DNAzyme domain for DNAzyme cleavage. An assistant DNA (A-DNA) probe containing aptamer and complementary domains was also employed to recognize protein and to induce DNA assembly. Once H-DNA and A-DNA recognize the same protein, H-DNA and A-DNA are in close proximity to each other. This induces DNA assembling for protein-triggered complex (Protein-Complex) with free DNAzyme domains. The free DNAzymes trigger the circular cleavage of molecular beacons for amplified signals. The assay is performed by fluorometry at an excitation wavelength of 980 nm and by collecting fluorescence at 545 nm. The platelet derived growth factor-BB (PDGF-BB) was accurately identified and selectively determined by this assay with a 22 pM detection limit (using the 3σ criterion). The responses for PDGF-BB is nearly 6-fold higher than for PDGF-AB, and 16-fold higher than PDGF-AA. This upconversion assay avoids any interference by the autofluorescence of biological fluids.

Schematic representation of the principle of the target-triggered DNA assembling probes mediated amplification strategy based on affinity binding for PDGF-BB. The UCNP probe is used for the quantitation of PDGF-BB with high selectivity.

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Acknowledgements

This work was supported by the Tianjin Natural Science Foundation (Grant 18JCZDJC98400), and National Natural Science Foundation of China (Grant 21245001).

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Authors and Affiliations

  1. Key Laboratory of Inorganic-Organic Hybrid Functional Materials Chemistry (Tianjin Normal University), Ministry of Education, Tianjin Key Laboratory of Structure and Performance for Functional Molecules, College of Chemistry, Tianjin Normal University, Tianjin, 300387, China

    Yue Liu & Ying Wang

  2. Department of Pharmacology, Tianjin Nankai Hospital, Tianjin, 300100, China

    Wei-Jun Liu

  3. State Grid Economic and Technological Research Institute Co. LTD, Beijing, 102209, China

    Jia Hu

  4. Department of Chemistry, Nankai University, Tianjin, 300071, China

    Yan Li

  5. Tianjin North Food Co. Ltd., Tianjin, 300350, China

    Li-Xia Zhao

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  1. Yue Liu
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  2. Wei-Jun Liu
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  4. Yan Li
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  5. Ying Wang
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Correspondence to Yue Liu or Ying Wang.

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Liu, Y., Liu, WJ., Hu, J. et al. Affinity binding-mediated fluorometric protein assay based on the use of target-triggered DNA assembling probes and aptamers labelled with upconversion nanoparticles: application to the determination of platelet derived growth factor-BB. Microchim Acta 187, 9 (2020). https://doi.org/10.1007/s00604-019-4024-2

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