Imaging and photobleach correction of Mero-CBD, sensor of endogenous Cdc42 activation

Louis Hodgson, Perihan Nalbant, Feimo Shen, Klaus Hahn

Research output: Contribution to journalReview article

39 Scopus citations

Abstract

This chapter details quantitative imaging of the Mero-CBD biosensor, which reports activation of endogenous Cdc42 in living cells. The procedures described are appropriate for imaging any biosensor that uses two different fluorophores on a single molecule, including FRET biosensors. Of particular interest is an algorithm to correct for fluorophore photobleaching, useful when quantitating activity changes over time. Specific topics include procedures and caveats in production of the Mero-CBD sensor, image acquisition, motion artifacts, shading correction, background subtraction, registration, and ratio imaging.

Original languageEnglish (US)
Article number12
Pages (from-to)140-156
Number of pages17
JournalMethods in enzymology
Volume406
DOIs
StatePublished - Jan 1 2006
Externally publishedYes

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ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology

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