Development of an aptamer beacon for detection of Interferon-Gamma

Nazgul Tuleuova, Caroline N. Jones, Jun Yan, Erlan Ramanculov, Yohei Yokobayashi, Alexander Revzin

Research output: Contribution to journalArticle

97 Citations (Scopus)

Abstract

Traditional antibody-based affinity sensing strategies employ multiple reagents and washing steps and are unsuitable for real-time detection of analyte binding. Aptamers, on the other hand, may be designed to monitor binding events directly, in real-time, without the need for secondary labels. The goal of the present study was to design an aptamer beacon for fluorescence resonance energy transfer (FRET)based detection of interferon-gamma (IFN-γ)-an important inflammatory cytokine. Variants of DNA aptamer modified with biotin moieties and spacers were immobilized on avidin-coated surfaces and characterized by surface plasmon resonance (SPR). The SPR studies showed that immobilization of aptamer via the 3' end resulted in the best binding IFN-γ (Kd = 3.44 nM). This optimal aptamer variant was then used to construct a beacon by hybridizing fluorophore-labeled aptamer with an antisense oligonucleotide strand carrying a quencher. SPR studies revealed that IFN-γ binding with an aptamer beacon occurred within 15 min of analyte introduction-suggesting dynamic replacement of the quencher-complementary strand by IFN-γ molecules. To further highlight biosensing applications, aptamer beacon molecules were immobilized inside microfluidic channels and challenged with varying concentration of analyte. Fluorescence microscopy revealed low fluorescence in the absence of analyte and high fluorescence after introduction of IFN-γ. Importantly, unlike traditional antibody-based immunoassays, the signal was observed directly upon binding of analyte without the need for multiple washing steps, The surface immobilized aptamer beacon had a linear range from 5 to 100 nM and a lower limit of detection of 5 nM IFN-γ. In conclusion, we designed a FRET-based aptamer beacon for monitoring of an inflammatory cytokine-IFNy. In the future, this biosensing strategy will be employed to monitor dynamics of cytokine production by the immune cells.

Original languageEnglish (US)
Pages (from-to)1851-1857
Number of pages7
JournalAnalytical Chemistry
Volume82
Issue number5
DOIs
StatePublished - Mar 1 2010
Externally publishedYes

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Interferon-gamma
Surface plasmon resonance
Cytokines
Washing
Fluorescence
Nucleotide Aptamers
Molecules
Avidin
Fluorophores
Antibodies
Antisense Oligonucleotides
Fluorescence microscopy
Biotin
Microfluidics
Labels
Monitoring
Fluorescence Resonance Energy Transfer

ASJC Scopus subject areas

  • Analytical Chemistry

Cite this

Tuleuova, N., Jones, C. N., Yan, J., Ramanculov, E., Yokobayashi, Y., & Revzin, A. (2010). Development of an aptamer beacon for detection of Interferon-Gamma. Analytical Chemistry, 82(5), 1851-1857. https://doi.org/10.1021/ac9025237

Development of an aptamer beacon for detection of Interferon-Gamma. / Tuleuova, Nazgul; Jones, Caroline N.; Yan, Jun; Ramanculov, Erlan; Yokobayashi, Yohei; Revzin, Alexander.

In: Analytical Chemistry, Vol. 82, No. 5, 01.03.2010, p. 1851-1857.

Research output: Contribution to journalArticle

Tuleuova, N, Jones, CN, Yan, J, Ramanculov, E, Yokobayashi, Y & Revzin, A 2010, 'Development of an aptamer beacon for detection of Interferon-Gamma', Analytical Chemistry, vol. 82, no. 5, pp. 1851-1857. https://doi.org/10.1021/ac9025237
Tuleuova N, Jones CN, Yan J, Ramanculov E, Yokobayashi Y, Revzin A. Development of an aptamer beacon for detection of Interferon-Gamma. Analytical Chemistry. 2010 Mar 1;82(5):1851-1857. https://doi.org/10.1021/ac9025237
Tuleuova, Nazgul ; Jones, Caroline N. ; Yan, Jun ; Ramanculov, Erlan ; Yokobayashi, Yohei ; Revzin, Alexander. / Development of an aptamer beacon for detection of Interferon-Gamma. In: Analytical Chemistry. 2010 ; Vol. 82, No. 5. pp. 1851-1857.
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