Ultrastructural analysis of myoblast fusion in Drosophila.
Myoblast fusion in Drosophila has become a powerful genetic system with which to unravel the mechanisms underlying cell fusion. The identification of important components of myoblast fusion by genetic analysis has led to a molecular pathway toward our understanding of this cellular process. In addition to the application of immunohistochemistry and live imaging techniques to visualize myoblast fusion at the light microscopic level, ultrastructural analysis using electron microscopy remains an indispensable tool to reveal fusion intermediates and specific membrane events at sites of fusion. In this chapter, we describe conventional chemical fixation and high-pressure freezing/freeze substitution methods for visualizing fusion intermediates during Drosophila myoblast fusion. Furthermore, we describe an immunoelectron microscopic method for localizing specific proteins relative to the fusion apparatus.
Zhang
Shiliang
S
Department of Molecular Biology and Genetics, The Johns Hopkins University School of Medicine, Baltimore, MD, USA.
Chen
Elizabeth H
EH
D000818:Animals; D002459:Cell Fusion / Q000379:methods*; D004331:Drosophila melanogaster / Q000166:cytology* / Q000196:embryology / Q000648:ultrastructure*; D004625:Embryo, Nonmammalian / Q000166:cytology / Q000648:ultrastructure; D005615:Freezing; D016253:Microscopy, Immunoelectron / Q000379:methods*; D032446:Myoblasts / Q000166:cytology* / Q000648:ultrastructure*; D016707:Tissue Fixation