Zebrafish embryo preparation for imaging in DaXi light-sheet microscope .md

Zebrafish embryo preparation for imaging in DaXi light-sheet microscope

This protocol outlines the steps for preparing zebrafish embryos for imaging in a DaXi light-sheet microscope. The protocol involves dechorionation, embedding in low-gelling temperature agarose, positioning embryos for imaging, removing the agarose surrounding the tail, and setting up time-lapses with tricaine to prevent embryo movement.

Materials:

• Glass-bottom cell culture dish (35mm Cell Culture Dish with Glass Bottom 20mm – Stellar Scientific)
• Low-gelling temperature agarose (Sigma, A0701)
• Custom-made capillary
• Dissection knife
• Forceps
• Embryo-medium solution
• Tricaine (0.016%)

Protocol:

1. Dechorionate embryos in a glass-bottom cell culture dish.
2. Gently transfer embryos into a 0.7% solution of low-gelling temperature agarose.
3. Position embryos at the correct position and angle for imaging using a custom-made capillary.
4. Once the agarose solidifies, use a dissection knife and forceps to remove the agarose surrounding the tail to permit full development and tail elongation.
5. Set up time lapses with a gentle flow of fresh and filtered embryo-medium solution with 0.016% tricaine to prevent embryo movement (after 15 hpf).