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Live imaging of epidermal morphogenesis during the development of the adult abdominal epidermis of Drosophila

Abstract

During larval stages of Drosophila development, the abdominal epidermis is composed of histoblasts (adult precursors) and larval epidermal cells (LECs). During metamorphosis, histoblasts proliferate and colonize the territories occupied by the LECs, which die and become engulfed by macrophages. This morphogenetic process is an excellent model for in vivo analysis of epithelial migration, cell division, cell death, patterning and differentiation. Here, we describe a protocol for time-lapse recording of the developing epidermis during metamorphosis. The protocol describes the removal of the pupal case (which acts as an opaque barrier to effective imaging) and mounting and imaging of specimens of different stages so that normal developmental processes are preserved. This method enables high-resolution studies over long time periods using fluorescent markers and confocal microscopy. The protocol requires 1 h for pupal dissection and mounting and, depending on the stages and genotypes to be analyzed, several more hours for preprocessing and aging and developmental staging of flies and pupae.

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Figure 1: Visualization of histoblasts in early and late pupae.
Figure 2: Orientation of live pupae on the imaging stage.
Figure 3: Dissection procedure for late pupae.
Figure 4: Timeline of adult Drosophila abdominal epithelia development.
Figure 5: Histoblast imaging in early pupae.
Figure 6: Histoblast imaging in late pupae.

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Acknowledgements

We thank Andrea Brand, Hiroki Oda, Roger Karess, Pernille Rorth and Elena Rebollo for fly stocks, Elena Rebollo for advice on setting the imaging acquisition, and Hector Herranz, lab members, reviewers and the NP editorial staff for help in improving the manuscript. Special thanks to Ashley Bruce for letting us use her equipment. N.N. holds an FPU studentship from the Spanish Ministery of Education, and E.M.-B. is supported by grants from the DGICYT (Spain) and the EU (WOUND STREP project).

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Correspondence to Enrique Martín-Blanco.

Supplementary information

Supplementary Video 1

Dissection of late pupae for live imaging (MOV 2064 kb)

Supplementary Video 2

Live imaging of early pupae (MOV 1579 kb)

Supplementary Video 3

Live imaging of late pupae (MOV 3660 kb)

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Ninov, N., Martín-Blanco, E. Live imaging of epidermal morphogenesis during the development of the adult abdominal epidermis of Drosophila. Nat Protoc 2, 3074–3080 (2007). https://doi.org/10.1038/nprot.2007.417

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