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Live-cell Measurement of Odorant Receptor Activation Using a Real-time cAMP Assay.

Publication ,  Journal Article
Zhang, Y; Pan, Y; Matsunami, H; Zhuang, H
Published in: J Vis Exp
October 2, 2017

The enormous sizes of the mammalian odorant receptor (OR) families present difficulties to find their cognate ligands among numerous volatile chemicals. To efficiently and accurately deorphanize ORs, we combine the use of a heterologous cell line to express mammalian ORs and a genetically modified biosensor plasmid to measure cAMP production downstream of OR activation in real time. This assay can be used to screen odorants against ORs and vice versa. Positive odorant-receptor interactions from the screens can be subsequently confirmed by testing against various odor concentrations, generating concentration-response curves. Here we used this method to perform a high-throughput screening of an odorous compound against a human OR library expressed in Hana3A cells and confirmed that the positively-responding receptor is the cognate receptor for the compound of interest. We found this high-throughput detection method to be efficient and reliable in assessing OR activation and our data provide an example of its potential use in OR functional studies.

Duke Scholars

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Published In

J Vis Exp

DOI

EISSN

1940-087X

Publication Date

October 2, 2017

Issue

128

Location

United States

Related Subject Headings

  • Transfection
  • Smell
  • Response Elements
  • Receptors, Odorant
  • Luminescent Measurements
  • Humans
  • High-Throughput Screening Assays
  • Cycloparaffins
  • Cyclic AMP
  • Computer Systems
 

Citation

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Zhang, Y., Pan, Y., Matsunami, H., & Zhuang, H. (2017). Live-cell Measurement of Odorant Receptor Activation Using a Real-time cAMP Assay. J Vis Exp, (128). https://doi.org/10.3791/55831
Zhang, Yuetian, Yi Pan, Hiroaki Matsunami, and Hanyi Zhuang. “Live-cell Measurement of Odorant Receptor Activation Using a Real-time cAMP Assay.J Vis Exp, no. 128 (October 2, 2017). https://doi.org/10.3791/55831.
Zhang Y, Pan Y, Matsunami H, Zhuang H. Live-cell Measurement of Odorant Receptor Activation Using a Real-time cAMP Assay. J Vis Exp. 2017 Oct 2;(128).
Zhang, Yuetian, et al. “Live-cell Measurement of Odorant Receptor Activation Using a Real-time cAMP Assay.J Vis Exp, no. 128, Oct. 2017. Pubmed, doi:10.3791/55831.
Zhang Y, Pan Y, Matsunami H, Zhuang H. Live-cell Measurement of Odorant Receptor Activation Using a Real-time cAMP Assay. J Vis Exp. 2017 Oct 2;(128).

Published In

J Vis Exp

DOI

EISSN

1940-087X

Publication Date

October 2, 2017

Issue

128

Location

United States

Related Subject Headings

  • Transfection
  • Smell
  • Response Elements
  • Receptors, Odorant
  • Luminescent Measurements
  • Humans
  • High-Throughput Screening Assays
  • Cycloparaffins
  • Cyclic AMP
  • Computer Systems