Allosteric aptamers controlling a signal amplification cascade allow visual detection of molecules at picomolar concentrations

Biochemistry. 2006 Feb 21;45(7):2461-6. doi: 10.1021/bi052106i.

Abstract

A broadly applicable homogeneous detection system has been developed. It utilizes components of the blood coagulation cascade in the presence of polystyrene microspheres (MS) as a signal amplifier. Russell's viper venom factor X activator (RVV-X) triggers the cascade, which results in an eye-visible phase transition (precipitation) of MS bound to clotted fibrin. An allosteric RNA aptamer, RNA132, with affinity for RVV-X and human vascular endothelial growth factor (VEGF(165)) was created. RNA132 inhibits enzymatic activity of RVV-X. The effector molecule, VEGF(165), reverses the inhibitory activity of RNA132 on RVV-X and restores its enzymatic activity, thus, triggering the cascade and enabling the phase transition. As few as 5 fmol of VEGF(165) could be detected by the naked eye within an hour. Similar results were obtained for another allosteric aptamer modulated by a protein tyrosine phosphatase. The assay is instrumentation-free for both processing and readout and can be modified to detect molecules to which aptamers can be obtained.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Allosteric Regulation*
  • Aptamers, Nucleotide / pharmacology*
  • Base Sequence
  • Blood Coagulation / physiology*
  • Humans
  • Kinetics
  • Metalloendopeptidases / metabolism*
  • Microspheres
  • Molecular Sequence Data
  • Nanotechnology / methods
  • Signal Transduction*
  • Vascular Endothelial Growth Factor A / metabolism

Substances

  • Aptamers, Nucleotide
  • VEGFA protein, human
  • Vascular Endothelial Growth Factor A
  • Metalloendopeptidases
  • russellysin