Welcome to the laboratory of Dr.Michael Kalen. I'm Kim Gard. Today we'll demonstrate a skin transplantation model using the allogeneic BC to black six strain combination as one of the most stringent and least technically challenging models of tissue graft rejection.
Skin transplantation is a standard technique employed to assay allogeneic responses and is commonly used to assess efficiency of various tolerance induction strategies. So let's begin. This procedure starts with harvesting the skin from our BAB seed donor mouse.
First euthanize the donor in accordance with Iacuc SOP. Now wet hair back at base with alcohol, harvest skin by removing the ear as close to the base as possible and place on ice. Next, split each ear year discarding the collagenous ventral flap and float the tissue on ice.
Now the donor skin is ready for transplantation. The transplant procedure begins by anesthetizing the recipient animal using approved procedures. Once the recipient is sufficiently anesthetized, remove the animal from the chamber and give bup andnext IP for pain relief.
Place the recipient in a lateral position and assure the animal is fully sedated by performing toe. Pinch wet hair with alcohol and shave. Once recipient is cleanly shaven, make approximately one to two centimeter incision.
Place donor scan atop the incision And remove excess fluid with sterile Q-tip. Now carefully trim away the excess donor skin wrap, animal and bandage. Make sure bandage is not too tight by assessing, breathing, and then check that the arms are freely mobile.
Now secure the bandage with a single suture. Place the mouse in a clean cage with a heat lamp until the animal has recovered for the first few days. Following surgery, you should monitor the animal a couple of times a day for signs of distress and give bup necks as needed for pain.
Five to seven days Following surgery, the bandages are removed. I'll be demonstrating bandage removal in a recipient from day six post-transplant. Once the mouse is fully anesthetized, remove the suture and using a blunt nose scissors cut through the bandage and remove gently, making sure not to disrupt the transplant.
Return the animal to a clean cage and monitor daily for signs of rejection. Depending on the strain combination you choose, most skin transplants are rejected, which is usually defined as 80%necrosis by day eight to 12. This mouse is day eight post-op and has received bilateral transplants.
On the left is a control syngeneic black six transplant, which will not be rejected. On the right is the allogeneic BAB sea transplant that will be rejected. Here's another date transplant that is in the process of being rejected.
Finally, a different date transplant that is almost completely rejected. So these three examples demonstrate the normal variation in kinetics of skin transplant rejection. In the preceding experiment, we've shown you how to perform a mirroring skin transplant.
This model is a quick and easy method of monitoring allogeneic responses and has a wide range of applications. For example, skin transplantation can be used to assay the efficiency of various immunosuppressive agents blocking antibodies and tolerance induction protocols. So that's it.
Thanks for watching and good luck with your experiments.
