Live Imaging of the Drosophila Pupal Eye Using Fluorescence Microscopy

Begin with a transgenic Drosophila pupa with an exposed eye.

Place the pupa on a jelly bead within a hydrated paper frame, surrounded by a jelly ring.

The pupa’s eye contains GFP-tagged junction proteins that highlight cell boundaries in the developing eye’s neuroepithelium.

Gently press a coverslip onto the pupal eye region.

Under a fluorescent microscope, GFP-tagged junctions fluoresce, displaying the neuroepithelium organization.

Take images at regular intervals across different depths and process the images to enhance contrast and minimize background noise.

Align these images with increasing time intervals to track the developing neuroepithelium.

Initially, primary cells form boundaries around photoreceptor cell clusters, while interommatidial cells or ICs arrange into a single layer.

Later, the ICs differentiate into secondary and tertiary cells near photoreceptors.

Finally, excess ICs are removed via programmed cell death, forming a hexagonal lattice that structures the organized eye.

Using forceps, carefully lift and remove the operculum to expose the pupal head. Next, tear along the side of the pupal case to expose the area around one eye. Gently remove the pupa from the adhesive t