Journal of Fluorescence

, Volume 24, Issue 6, pp 1581–1591 | Cite as

Novel Photochrome Aptamer Switch Assay (PHASA) for Adaptive Binding to Aptamers

  • Vladislav Papper
  • Oleksandr Pokholenko
  • Yuanyuan Wu
  • Yubin Zhou
  • Ping Jianfeng
  • Terry W. J. Steele
  • Robert S. Marks
ORIGINAL PAPER

Abstract

A novel Photochrome-Aptamer Switch Assay (PHASA) for the detection and quantification of small environmentally important molecules such as toxins, explosives, drugs and pollutants, which are difficult to detect using antibodies-based assays with high sensitivity and specificity, has been developed. The assay is based on the conjugation of a particular stilbene-analyte derivative to any aptamer of interest. A unique feature of the stilbene molecule is its reporting power via trans-cis photoisomerisation (from fluorescent trans-isomer to non-fluorescent cis-isomer) upon irradiation with the excitation light. The resulting fluorescence decay rate for the trans-isomer of the stilbene-analyte depends on viscosity and spatial freedom to rotate in the surrounding medium and can be used to indicate the presence of the analyte. Quantification of the assay is achieved by calibration of the fluorescence decay rate for the amount of the tested analyte. Two different formats of PHASA have been recently developed: direct conjugation and adaptive binding. New stilbene-maleimide derivatives used in the adaptive binding format have been prepared and characterised. They demonstrate effective binding to the model thiol compound and to the thiolated Malachite Green aptamer.

Keywords

Stilbene Aptamer Fluorescence Photoisomerisation Maleimide 

Abbreviations

2b

4-methoxy-4′-stilbene maleimide

2a

4-N,N′-dimethylamino-4′-stilbene maleimide

MG

Malachite green

DMSO

Dimethyl sulfoxide

DMF

Dimethyl formamide

MCH

6-mercaptohexanol

EX

Fluorescence excitation

EM

Fluorescence emission

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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  • Vladislav Papper
    • 1
  • Oleksandr Pokholenko
    • 1
  • Yuanyuan Wu
    • 1
  • Yubin Zhou
    • 1
  • Ping Jianfeng
    • 1
  • Terry W. J. Steele
    • 1
  • Robert S. Marks
    • 2
  1. 1.School of Materials Science and EngineeringNanyang Technological UniversitySingaporeSingapore
  2. 2.Department of Biotechnology Engineering, National Institute for Biotechnology in the Negev, Ilse Kats Institute for Nanoscale Science and TechnologyBen Gurion University of the NegevBeer ShevaIsrael

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