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Colorimetric enzyme-linked aptamer assay utilizing hybridization chain reaction for determination of bovine pregnancy-associated glycoproteins

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Abstract

DNA aptamers that bind to bovine pregnancy-associated glycoproteins (bPAGs) were selected by the systematic evolution of ligands by exponential enrichment (SELEX) procedure coupled to surface plasmon resonance (SPR) and high-throughput sequencing (HTS) technology. After seven rounds of selection using carboxylated magnetic beads (MB) coated with bovine pregnancy-associated glycoproteins 9 (bPAG9) and bovine serum albumin (BSA) as target and counter targets, respectively, two aptamers designated as A1 and A24 showed high affinities to bPAG9 (Kd = 1.04 and 2.5 nM). Moreover, the specificity was determined by testing the non-targets bPAG4, bPAG6, bPAG16, BSA, and ovalbumin (OVA). Results showed that two aptamers demonstrated broad group specificity to bPAG family. Subsequently, a colorimetric sandwich enzyme-linked aptamer assay was developed for ultrasensitive detection of bPAG9 based on hybridization chain reaction (HCR) amplification strategy. The method exhibited a broad determination from 0.134 to 134 ng/mL with a detection limit of 0.037 ng/mL. The method has been successfully applied to determine bPAGs in real samples. The results demonstrate that the developed aptamers could be used as promising molecular probes for the development of pregnancy diagnostic tools.

In this study, we first selected aptamers against bovine pregnancy-associated glycoproteins (bPAGs) as molecular recognition elements and then developed a colorimetric enzyme-linked aptamer assay utilizing hybridization chain reaction to detect bPAGs in the serum.The GA can't be deleted, the modified GA can not upload. So themodified GA and figures will be send to CorrAdmin3@spi-global.com

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Funding

This work was financially supported by the National Natural Science Foundation of China (31860647).

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Authors and Affiliations

  1. College of Life Science and Technology, Yangtze Normal University, Chongqing, 408100, People’s Republic of China

    Chunxia Lu

  2. Institute of Animal Husbandry and Veterinary Science, Xinjiang Academy of Agriculture and Reclamation Science, Shihezi, 832000, People’s Republic of China

    Changbin Liu & Guoqing Shi

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  1. Chunxia Lu
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  2. Changbin Liu
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  3. Guoqing Shi
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Correspondence to Changbin Liu.

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All experimental animal procedures were performed in compliance with the China Code of Practice Animals for the Care and Use of Animals for Scientific Purposes. The animal welfare was approved by Animal Experimental Ethical Review Form of the First Affiliated Hospital of Medical College, Shihezi University.

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The authors declare that they have no competing of interests.

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Lu, C., Liu, C. & Shi, G. Colorimetric enzyme-linked aptamer assay utilizing hybridization chain reaction for determination of bovine pregnancy-associated glycoproteins. Microchim Acta 187, 316 (2020). https://doi.org/10.1007/s00604-020-04301-y

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  • DOI: https://doi.org/10.1007/s00604-020-04301-y

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