Multicolor Imaging of Bifacial Activities of Estrogens

  • Sung-Bae KimEmail author
  • Yoshio Umezawa
Part of the Methods in Molecular Biology book series (MIMB, volume 1461)


The present protocol introduces multicolor imaging of bifacial activities of an estrogen. For the multicolor imaging, the authors fabricated two single-chain probes emitting green or red bioluminescence (named Simer-G and -R, respectively) from click beetle luciferase (CBLuc) green and red: Simer-R consists of the ligand binding domain of estrogen receptor (ER LBD) and the Src homology-2 (SH2) domain of Src, which are sandwiched between split-CBLuc red (CBLuc-R). On the other hand, Simer-G emitting red light consists of the ER LBD and a common consensus sequence of coactivators (LXXLL motif), which are inserted between split-CBLuc green (CBLuc-G). This probe set creates fingerprinting spectra from the characteristic green and red bioluminescence in response to agonistic and antagonistic activities of a ligand of interest. The present protocol further provides a unique methodology to calculate characteristic estrogenicity scores of various ligands from the spectra.

Key words

Bioluminescence Multicolor imaging Luciferase Single-chain probe Estrogen receptor Click beetle 



This work was supported by grants from Japan Society for the Promotion of Science (JSPS), grant numbers 26288088, 15KK0029, and 16K14051.


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Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  1. 1.Research Institute for Environmental Management TechnologyNational Institute of Advanced Industrial Science and Technology (AIST)TsukubaJapan
  2. 2.Department of Chemistry, School of ScienceThe University of TokyoBunkyo-ku, TokyoJapan

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