After completing the primary euthanasia, check the loss of pain response by pinching the foot. Place the frog on its back. Pin down each limb.
Cut through the midline of the skin, then cut laterally to create two skin flaps. Grab the linea alba with forceps, and pull it away from the coelomic cavity. Cut through the musculature.
Cut or pin all the flaps out of the way. Cut through the coracoid bones with the help of dissection scissors and remove any excess tissue to gain better access to the heart. Next, gently shift the stomach of the dissected Xenopus to the top of the left liver lobe.
Gently grasp a lung tip using tissue forceps and pull it outside the coelomic cavity, then pin the lung through the tip. Pull the thin pericardium tight with tissue forceps. Using the tip of a pair of iridectomy scissors, gently perforate the pericardium without cutting any underlying tissue.
Peel the pericardium away from the heart chambers. Gently grasp the ventricle apex with forceps. Without perforating the ventricle, insert the needle through the closure of the forceps into the ventricle chamber.
Clamp the tissue forceps in place using a needle holder or hemostat. Start the flow of the pump at approximately five milliliters per minute. This will engorge the heart chambers and the arterial trunk.
Use scissors to make a nick in the right auricle. Continue the pump flow until the vasculature of the stomach blanches. Lance the left heart auricle.
Increase the flow rate of the pump to 10 milliliters per minute. Using a transfer pipette, rinse the coelomic cavity and perfusion media. If the organs shift during rinsing, gently shift them back to maintain visibility of the stomach and lungs.
To help visualize the perfusate flowing out of the heart, the auricles may further be reduced. Keep the needle in place until the stomach vasculature and lungs have blanched and the perfusate flowing from the heart appears clear. Successful perfusion was observed in all tissues except the liver, making them distinctly lighter and less saturated with blood.
This is most visible in albinos.
