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Related Concept Videos

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Labeling DNA Probes

DNA probes are fragments of DNA labeled with a reporter tag to enable their detection or purification. The resulting labeled DNA probes can then hybridize to target nucleic acid sequences through complementary base-pairing, and may be used to recover or identify these regions.
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Split Hybridization Probe Utilizing a DNA Fluorescent Light-up Aptamer as a Signal Reporter for Sequence-Specific Nucleic Acid Analysis
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Enzyme-linked small-molecule detection using split aptamer ligation.

Ashwani K Sharma1, Alexandra D Kent, Jennifer M Heemstra

  • 1Department of Chemistry, University of Utah, Salt Lake City, Utah 84112, United States.

Analytical Chemistry
|June 22, 2012
PubMed
Summary
This summary is machine-generated.

A new aptamer-based assay detects cocaine using split DNA strands that assemble only in its presence. This method offers a two-order-of-magnitude improvement in detection limits for cocaine in human serum.

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A Method for Selecting Structure-switching Aptamers Applied to a Colorimetric Gold Nanoparticle Assay

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Area of Science:

  • Biochemistry
  • Molecular Biology
  • Analytical Chemistry

Background:

  • The enzyme-linked immunosorbent assay (ELISA) is a common method for detecting molecules.
  • Aptamers are short DNA or RNA sequences that can bind to specific targets.
  • Split aptamers offer a unique approach for molecular detection by requiring target-induced assembly.

Purpose of the Study:

  • To develop an aptamer-based assay as an alternative to traditional ELISA for detecting small molecules.
  • To create a cocaine detection assay utilizing the cocaine split aptamer system.
  • To improve the sensitivity and utility of split aptamer-based sensors.

Main Methods:

  • Utilizing a cocaine split aptamer composed of two DNA strands that assemble upon cocaine binding.
  • Immobilizing one aptamer fragment on a microplate and introducing the second fragment in the test sample.
  • Employing click chemistry (azide-cyclooctyne ligation) for covalent biotinylation upon aptamer assembly.
  • Developing a colorimetric readout via streptavidin-horseradish peroxidase conjugation.

Main Results:

  • Demonstrated detection of cocaine in buffer across a concentration range of 100 nM to 100 μM.
  • Successfully detected cocaine in human blood serum within the concentration range of 1 μM to 100 μM.
  • Achieved a detection limit of 1 μM in serum, representing a significant improvement over existing split aptamer sensors.

Conclusions:

  • The developed aptamer-based assay effectively detects cocaine with high sensitivity.
  • Covalently trapping split aptamer assembly events enhances sensor performance.
  • This assay shows promise as a sensitive and specific method for small molecule detection in biological samples.