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Bio Protoc. 2017 Jul 5;7(13). pii: e2380. doi: 10.21769/BioProtoc.2380.

Ex vivo Ooplasmic Extract from Developing Drosophila Oocytes for Quantitative TIRF Microscopy Analysis.

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1
European Molecular Biology Laboratory (EMBL), Developmental Biology Unit, Heidelberg, Meyerhofstrasse 1, D-69117, Germany.

Abstract

Understanding the dynamic behavior and the continuously changing composition of macromolecular complexes, subcellular structures and organelles is one of areas of active research in both cell and developmental biology, as these changes directly relate to function and subsequently to the development and homeostasis of the organism. Here, we demonstrate the use of the developing Drosophila oocyte to study dynamics of messenger ribonucleoprotein complexes (mRNPs) with high spatiotemporal resolution. The combination of Drosophila genetics with total internal reflection (TIRF) microscopy, image processing and data analysis gives insight into mRNP motility and composition dynamics with unprecedented precision.

KEYWORDS:

Intracellular motility; Oocyte; Ooplasmic extract; TIRF microscopy; ex vivo assay

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