0 views • 3:27 min • April 30th, 2023
- Start with a Petri dish containing embryos in E3 medium. Add tricaine to anesthetize the embryos and phenylthiouria, PTU solution, to inhibit pigment formation. With the help of a dissecting microscope, dechorionate the embryos.
Use a glass pipette to transfer one of the dechorionated embryos to a small Petri dish with a shallow glass bottom well in the center and remove the excess medium. Next, place an agarose solution at an optimum concentration to minimize embryo distortion and motility into a microfuge tube and heat it. Bring down the temperature to 30 degrees Celsius to avoid damaging the embryo due to heat.
Pour the first layer of agarose over the embryo, completely filling the shallow well. Place another cover glass over the well and add 1% agarose on top of that. The second layer holds the cover glass in place.
Once the agarose solidifies, fill the Petri dish with the E3 medium containing tricaine. This is the third layer and keeps the agarose and embryo hydrated. In the following protocol, we will perform layered agar mounting of zebrafish embryos for extended time lapse imaging.
- Just before mounting, heat the agarose solution to 65 degrees Celsius and then let it coo
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