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Rab GTPases pp 181-186 | Cite as

3D Time-Lapse Analysis of Rab11/FIP5 Complex: Spatiotemporal Dynamics During Apical Lumen Formation

  • Anthony Mangan
  • Rytis PrekerisEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1298)

Abstract

Fluorescent imaging of fixed cells grown in two-dimensional (2D) cultures is one of the most widely used techniques for observing protein localization and distribution within cells. Although this technique can also be applied to polarized epithelial cells that form three-dimensional (3D) cysts when grown in a Matrigel matrix suspension, there are still significant limitations in imaging cells fixed at a particular point in time. Here, we describe the use of 3D time-lapse imaging of live cells to observe the dynamics of apical membrane initiation site (AMIS) formation and lumen expansion in polarized epithelial cells.

Key words

Apical lumen Time-lapse microscopy Matrigel 3D tissue culture Epithelial cell polarity 

Notes

Acknowledgement

This work was supported by NIH grant R01-DK064380 to R.P. and NIH Pre-doctoral Training Program T32-GM08730.

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Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  1. 1.Department of Cell and Developmental Biology, School of MedicineUniversity of Colorado DenverAuroraUSA

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