Live Imaging of Drosophila Embryos: Quantifying Protein Numbers and Dynamics at Subcellular Locations

  • Daryl J. V. David
  • Melanie A. McGill
  • R. F. Andrew McKinley
  • Tony J. C. HarrisEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 839)


Live imaging is critical for understanding the structure and activities of protein interaction networks in cells. By tagging proteins of interest with fluorescent proteins, such as green fluorescent protein (GFP), their localization in cells can be determined and correlated with cellular activities. This can be extended into developmental systems such as Drosophila to understand the molecular and cellular bases of development. In this chapter, we review sample preparation techniques and basic imaging considerations for Drosophila embryos. We then discuss how these techniques can be extended to count absolute protein numbers at specific subcellular locations, and determine their dynamics using fluorescence recovery after photobleaching (FRAP). These techniques can help reveal the structure and dynamics of protein complexes in live cells.

Key words

Drosophila Live imaging Protein counting Fluorescence recovery after photobleaching Adherens junctions Epithelia 



Work in our lab is supported by a CIHR operating grant and an NSERC operating grant. A. McKinley holds an Ontario Graduate Scholarship in Science and Technology. T. Harris holds a Tier 2 Canada Research Chair.


  1. 1.
    McGill MA, McKinley RF, Harris TJ (2009) Independent cadherin-catenin and Bazooka clusters interact to assemble adherens junctions. J Cell Biol. 185:787–796.PubMedCrossRefGoogle Scholar
  2. 2.
    Pope KL, Harris TJ (2008) Control of cell flattening and junctional remodeling during squamous epithelial morphogenesis in Drosophila. Development. 135:2227–2238.PubMedCrossRefGoogle Scholar
  3. 3.
    David DJ, Tishkina A, Harris TJ (2010) The PAR complex regulates pulsed actomyosin contractions during amnioserosa apical constriction in Drosophila. Development. 137:1645–1655.PubMedCrossRefGoogle Scholar
  4. 4.
    Wu JQ, Pollard TD (2005) Counting cytokinesis proteins globally and locally in fission yeast. Science. 310: 310–314.PubMedCrossRefGoogle Scholar
  5. 5.
    Wu JQ, McCormick CD, Pollard TD (2008) Chapter 9: Counting proteins in living cells by quantitative fluorescence microscopy with internal standards. Methods Cell Biol. 89:253–273.PubMedCrossRefGoogle Scholar
  6. 6.
    Fowlkes CC, Hendriks CL, Keranen, SV et al (2008) A quantitative spatiotemporal atlas of gene expression in the Drosophila blastoderm. Cell. 133:364–374.PubMedCrossRefGoogle Scholar
  7. 7.
    Lippincott-Schwartz J, Snapp E, Kenworthy A (2001) Studying protein dynamics in living cells. Nat Rev Mol Cell Biol. 2:444–456.PubMedCrossRefGoogle Scholar
  8. 8.
    Lippincott-Schwartz J, Altan-Bonnet N, Patterson GH (2003) Photobleaching and ­photoactivation: following protein dynamics in ­living cells. Nat Cell Biol. Suppl: 5:S7–14.Google Scholar

Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  • Daryl J. V. David
    • 1
  • Melanie A. McGill
    • 1
  • R. F. Andrew McKinley
    • 1
  • Tony J. C. Harris
    • 1
    Email author
  1. 1.Department of Cell & Systems BiologyUniversity of TorontoTorontoCanada

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