Chambers for Culturing and Immobilizing <i>Xenopus</i> Embryos and Organotypic Explants for Live Imaging.

Abstract

Live imaging of <i>Xenopus</i> embryos and organotypic explants can be challenging because of their large size and slippery nature. This protocol covers the preparation of special chambers for immobilizing <i>Xenopus</i> embryos and embryonic explants for live-cell and tissue imaging. The opaque nature of <i>Xenopus</i> embryonic tissues enables simple bright-field imaging techniques for tracking surface movements across large regions. Such surface imaging of embryos or organotypic explants can directly reveal cell behaviors, obviating the need for complex postprocessing commonly required to extract this data from 3D confocal or light-sheet observations of more transparent embryos. Furthermore, <i>Xenopus</i> embryos may be filled with light-absorbing pigment granules and light-scattering yolk platelets, but these limitations are offset by the utilitarian nature of <i>Xenopus</i> organotypic explants that expose and stabilize large embryonic cells in a nearly native context for high-resolution live-cell imaging. Additionally, whole embryos can be stabilized for long-term bright-field and confocal microscopy. Simple explants can be prepared using a single cell type, and organotypic explants can be prepared in which multiple tissue types are dissected while retaining native tissue-tissue interactions. These preparations enable both in-toto imaging of tissue dynamics and super-resolution imaging of protein dynamics within individual cells. We present detailed protocols for these methods together with references to applications.