1 2 3 4 5 6 7

Procedure:
1. Obtain axolotl embryos. For this experiment, we will be using wild type and albino axolotl embryos at stage 21 in their staging series.
2. Prepare the axolotl embryos as follows:
a. manually dejelly embryo of appropriate stage and keep in
HEPES-buffered Modified Steinberg's solution (HBSt) containing antibiotics until use. For microsurgery, also remove membrane around embryo.
b. place embryos into operating dish containing 1 x HBSt and antibiotics
c. use clean technique; dip tools, pipettes, and glass bridges in 70% EtOH and then into sterile HBSt before using
3. Start
microsurgery on your donor embryo. Locate the gill organ field which is in the first anterior third of the embryo (toward head) and right above the small crest or indention closer to the ventral side (Rugh, 1962) . Isolate the gill organ field (or the region that will become the gill organ field) by cutting around it and detaching that specific piece of tissue from the embryo.
4. Start transplantation on your recipient embryo. Cut a small slit in a region close to the gill organ field on the recipient. Gently slip donor gill organ field into slit and be sure to secure the graft in place. It is critical to try to get the graft as far under the cut as possible to ensure that the recipient will begin to incorporate the new tissue into its development process.


Figure 1:

A. A 21 stage albino axolotl embryo with a transplanted wild type axolotl graft (supposedly the gill field). The picture was taken immediately after microsurgery.

B. A 21 stage wild type axolotl embryo with a transplanted albino axolotl graft. The picture was taken immediately after microsurgery

 

© Cebra-Thomas 2001

Last Modified: 5 August 2001


[Lab Protocols | Students | Cebra-Thomas | Course | Links ]