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Updated: Sep 26, 2025

Aptamer-Based Target Detection Facilitated by a 3-Stage G-Quadruplex Isothermal Exponential Amplification Reaction
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Systematically Modulating Aptamer Affinity and Specificity by Guanosine-to-Inosine Substitution.

Brea A Manuel1, Sierra A Sterling1, Aimee A Sanford1

  • 1Department of Chemistry, Emory University, Atlanta, Georgia 30322, United States.

Analytical Chemistry
|April 18, 2022
PubMed
Summary

Researchers developed a new method to tune aptamer binding affinity for biosensing by substituting guanosine with inosine. This approach offers a flexible way to create aptamers with desired detection ranges and specificities.

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Area of Science:

  • Biotechnology
  • Molecular Biology
  • Biosensing

Background:

  • Aptamers are crucial for small molecule detection due to their specificity, stability, and cost-effectiveness.
  • A key challenge in aptamer-based sensing is matching binding affinity to the target analyte concentration range.
  • Existing affinity modulation methods often cause significant, undesirable changes in aptamer folding.

Purpose of the Study:

  • To introduce a novel, less perturbative method for modulating aptamer binding affinity.
  • To explore the use of guanosine-to-inosine substitutions for tuning aptamer affinity and specificity.
  • To generate aptamers with a range of binding affinities for diverse biosensing applications.

Main Methods:

  • Systematic substitution of guanosine with inosine in the guanosine-to-inosine sequence of a known aptamer.
  • Utilized the cocaine binding aptamer as a model system for these modifications.
  • Assessed binding affinity, specificity, and folding stability (via melting temperature) of modified aptamers.

Main Results:

  • Generated aptamer sequences with a broad range of binding affinities, from 230 nM to 80 μM.
  • Demonstrated that guanosine-to-inosine substitutions can modulate aptamer specificity towards related analytes.
  • Observed expected impacts on aptamer folding stability due to the nucleotide substitutions.

Conclusions:

  • Guanosine-to-inosine substitution offers a convenient and reliable strategy for tuning aptamer binding affinity.
  • This method allows for the rapid generation of aptamers with tailored affinity and specificity for biosensing.
  • The approach provides a valuable tool for developing next-generation aptamer-based detection systems.