Evaluating cell-surface expression and measuring activation of mammalian odorant receptors in heterologous cells

Nat Protoc. 2008;3(9):1402-13. doi: 10.1038/nprot.2008.120.


A fundamental question in olfaction is which odorant receptors (ORs) are activated by a given odorant. A major roadblock to investigating odorant-OR relationships in mammals has been the inability to express ORs in heterologous cells suitable for screening active ligands for ORs. The discovery of the receptor-transporting protein family has facilitated the effective cell-surface expression of ORs in heterologous cells. The establishment of a robust heterologous expression system for mammalian ORs facilitates the high-throughput 'deorphanization' of these receptors by matching them to their cognate ligands. This protocol details the method used for evaluating the cell-surface expression and measuring the functional activation of ORs of transiently expressed mammalian ORs in HEK293T cells. The stages of OR cell-surface expression include cell culture preparation, transfer of cells, transfection, immunocytochemistry or flow cytometry, odorant stimulation and luciferase assay. This protocol can be completed in a period of 3 d from the transfer of cells to cell-surface expression detection and/or measurement of functional activation.

Publication types

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

MeSH terms

  • Cell Line
  • Flow Cytometry
  • Humans
  • Immunohistochemistry
  • Ligands
  • Luciferases
  • Molecular Biology / methods*
  • Receptors, Odorant / metabolism*
  • Signal Transduction / physiology
  • Smell / physiology*


  • Ligands
  • Receptors, Odorant
  • Luciferases