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Amplified detection of cocaine based on strand-displacement polymerization and fluorescence resonance energy transfer.

ABSTRACT: Cocaine is one of the most abused drugs in the United States and is potentially dangerous when consumed in excess. Its detection is thus important in many areas in the fight against drug trafficking. We have developed an amplified detection method for cocaine based on a strand-displacement polymerization reaction using aptamer recognition. The system mainly consists of a hairpin probe with Cy5 labeled on its 3' end, a primer with FAM labeled on its 5' end, and polymerase. The aptamer sequence is integrated into the 5'-section of the hairpin probe. The primer is designed complementary to the 3' end of the hairpin probe, which is also part of the hairpin stem region. The cocaine induced reaction cycle generates product for detection and thus for signal amplification. The detection limit of this method is 200 nM in about 16 min and the specificity of this approach is excellent. We believe that this strategy will be useful for the development of analytical schemes for a variety of aptamers for small molecules, metal ions, and proteins. This simple scheme employing the strand-displacement polymerization reaction may find wide application in forensic analysis, environmental monitoring, and clinical diagnostics.

SUBMITTER: Huang J

PROVIDER: S-EPMC3167194 | biostudies-literature | 2011 Oct

REPOSITORIES: biostudies-literature

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Amplified detection of cocaine based on strand-displacement polymerization and fluorescence resonance energy transfer.

Huang Jin J Chen Yan Y Yang Liu L Zhu Zhi Z Zhu Guizhi G Yang Xiaohai X Wang Kemin K Tan Weihong W

Biosensors & bioelectronics 20110601 1

Cocaine is one of the most abused drugs in the United States and is potentially dangerous when consumed in excess. Its detection is thus important in many areas in the fight against drug trafficking. We have developed an amplified detection method for cocaine based on a strand-displacement polymerization reaction using aptamer recognition. The system mainly consists of a hairpin probe with Cy5 labeled on its 3' end, a primer with FAM labeled on its 5' end, and polymerase. The aptamer sequence is ...[more]

PMID: 21802274

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Project description:Conjugated polydiacetylene (PDA) possessing stimuli-responsive properties has been intensively investigated for developing efficient sensors. We report here fluorescence resonance energy transfer (FRET) in liposomes synthesized using different molar ratios of dansyl-tagged diacetylene and diacetylene-carboxylic acid monomers. Photopolymerization of diacetylene resulted in cross-linked PDA liposomes. We used steady-state electronic absorption, emission, and fluorescence anisotropy (FA) analysis to characterize the thermal-induced FRET between dansyl fluorophores (donor) and PDA (acceptor). We found that the monomer ratio of acceptor to donor ( R ad) and length of linkers (functional part that connects dansyl fluorophores to the diacetylene group in the monomer) strongly affected FRET. For R ad = 10 000, the acceptor emission intensity was amplified by more than 18 times when the liposome solution was heated from 298 to 338 K. A decrease in R ad resulted in diminished acceptor emission amplification. This was primarily attributed to lower FRET efficiency between donors and acceptors and a higher background signal. We also found that the FRET amplification of PDA emissions after heating the solution was much higher when dansyl was linked to diacetylene through longer and flexible linkers than through shorter linkers. We attributed this to insertion of dansyl in the bilayer of the liposomes, which led to an increased dansyl quantum yield and a higher interaction of multiple acceptors with limited available donors. This was not the case for shorter and more rigid linkers where PDA amplification was much smaller. The present studies aim at enhancing our understanding of FRET between fluorophores and PDA-based conjugated liposomes. Furthermore, receptor tagged onto PDA liposomes can interact with ligands present on proteins, enzymes, and cells, which will produce emission sensing signal. Therefore, using the present approach, there exist opportunities for designing FRET-based highly sensitive and selective chemical and biochemical sensors.

Dataset Information

Amplified detection of cocaine based on strand-displacement polymerization and fluorescence resonance energy transfer.

Publications

Amplified detection of cocaine based on strand-displacement polymerization and fluorescence resonance energy transfer.

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