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Primer-Free Aptamer Selection Using A Random DNA Library
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Human thrombin detection through a sandwich aptamer microarray: interaction analysis in solution and in solid phase.

Alice Sosic1, Anna Meneghello, Erica Cretaio

  • 1Dipartimento di Scienze Farmaceutiche, University of Padova, via F. Marzolo 5, I-35131 Padova, Italy. alice.sosic@studenti.unipd.it

Sensors (Basel, Switzerland)
|December 14, 2011
PubMed
Summary

We developed an aptamer microarray for detecting human thrombin using two DNA aptamers. This biosensor targets different protein sites, enabling specific and sensitive detection.

Keywords:
aptamersandwich aptamer microarraythrombin

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Area of Science:

  • Biotechnology
  • Biosensor Development
  • Molecular Recognition

Background:

  • Human thrombin is a critical biomarker in various physiological and pathological processes.
  • Accurate detection of thrombin is essential for diagnosing and monitoring coagulation disorders and related conditions.
  • Existing detection methods may lack the sensitivity, specificity, or speed required for rapid clinical diagnostics.

Purpose of the Study:

  • To develop a novel aptamer-based microarray for sensitive and specific detection of human thrombin.
  • To utilize two distinct DNA aptamers targeting different exosites of human thrombin for enhanced recognition.
  • To optimize the sandwich aptamer microarray (SAM) assay for robust thrombin quantification.

Main Methods:

  • Development of an aptamer-based microarray platform.
  • Design and synthesis of two DNA aptamers (TBA1 and TBA2) with distinct binding sites on human thrombin.
  • Electrophoresis Mobility Shift Assay (EMSA) for validating aptamer-protein interactions in solution.
  • Optimization of the Sandwich Aptamer Microarray (SAM) assay protocol.
  • Evaluation of the specificity of the developed aptasensor.

Main Results:

  • Successful development of an aptamer-based microarray for human thrombin detection.
  • Demonstration that chemical modifications did not impede aptamer-thrombin recognition in solution.
  • Optimization of the SAM assay procedure for effective solid-phase detection.
  • Validation of the specificity of the aptasensor for human thrombin.

Conclusions:

  • The developed aptamer microarray offers a promising platform for sensitive and specific human thrombin detection.
  • The use of two aptamers targeting different exosites enhances the reliability of the biosensor.
  • The SAM assay provides a robust method for thrombin quantification in a microarray format.