A Sweet Spot for Molecular Diagnostics: Coupling Isothermal Amplification and Strand Exchange Circuits to Glucometers

Sci Rep. 2015 Jun 8:5:11039. doi: 10.1038/srep11039.

Abstract

Strand exchange nucleic acid circuitry can be used to transduce isothermal nucleic acid amplification products into signals that can be readable on an off-the-shelf glucometer. Loop-mediated isothermal amplification (LAMP) is limited by the accumulation of non-specific products, but nucleic acid circuitry can be used to probe and distinguish specific amplicons. By combining this high temperature isothermal amplification method with a thermostable invertase, we can directly transduce Middle-East respiratory syndrome coronavirus and Zaire Ebolavirus templates into glucose signals, with a sensitivity as low as 20-100 copies/μl, equating to atto-molar (or low zepto-mole). Virus from cell lysates and synthetic templates could be readily amplified and detected even in sputum or saliva. An OR gate that coordinately triggered on viral amplicons further guaranteed fail-safe virus detection. The method describes has potential for accelerating point-of-care applications, in that biological samples could be applied to a transducer that would then directly interface with an off-the-shelf, approved medical device.

Publication types

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

MeSH terms

  • Bacterial Proteins / chemistry*
  • Ebolavirus / chemistry*
  • Ebolavirus / genetics
  • Glucose / analysis*
  • Nucleic Acid Amplification Techniques*
  • Severe acute respiratory syndrome-related coronavirus / chemistry*
  • Severe acute respiratory syndrome-related coronavirus / genetics
  • beta-Fructofuranosidase / chemistry*

Substances

  • Bacterial Proteins
  • beta-Fructofuranosidase
  • Glucose