Orthotopic Homograft Tumor Transplantation: A Technique to Generate Pancreatic Tumor Mouse Models

- During orthotopic homograft tumor transplantation, tumor cells or tumor fragments from a cancerous donor get transplanted into a recipient belonging to the same species at an organ or site that matches the tumor histotype. Begin by taking an anesthetized donor mouse bearing a tumor, containing cells of pancreatic origin. Sterilize the area and surgically harvest the tumor. Wash the tissue to remove the blood and chop it into fragments. Next, place an anesthetized recipient mouse in a right lateral position and sterilize its abdominal area. Surgically expose the spleen.

Next, gently pull out a part of the pancreas from below the spleen. Place the tumor fragment on the pancreas and suture them together. Close the abdomen and allow the mouse to recover in a cage. Monitor the recipient mouse daily to observe the growth of the orthotopic pancreatic tumor. In the following protocol, we will show the orthotopic transplantation of pancreatic ductal adenocarcinoma homograft from a C57 black 6 donor mouse into the pancreas of a C57 black 6 recipient mouse.

- Monitor the bodyweight of C57 black 6 mice using a balance. Also, monitor the tumor volume of tumor-bearing donor mice by caliper measurement. Following euthanasia of the animal in a biohazard hood as per protocol, sterilize the skin around the tumor using iodophor swabs. After surgically removing the tumor as detailed in the text protocol, place the tumor in a Petri dish containing 20 milliliters of PBS that has been prechilled to four degrees Celsius. If there is contaminating blood, transfer the tumor into another Petri dish, and wash the tumor with PBS.

Cut the tumor in half. Remove any extra skin, vessels, calcification, and/or necrosis. Choose only intact pieces of tumor and place them into a sterile 50-milliliter centrifuge tube. Add 20 milliliters of PBS before transporting the tube to a separate animal room for pharmacology studies. To prepare for orthotopic implantation, fix the fully anesthetized mouse on an experiment board in the right lateral position. Disinfect the skin around the spleen with iodine and then deiodinate with 75% ethyl alcohol.

Find the medium point of the spleen and make a one-centimeter vertical incision on the abdomen to expose the spleen. Gently draw out a part of the pancreas tissue under the spleen using flat-tipped tweezers and suture a mouse homograft tumor piece from the seed mouse onto the pancreas of the recipient mouse by 9-0 absorbable surgical suture. Close the abdomen with a 6-0 silk suture by double seam. Achieve homeostasis by compression.

After finishing tumor implantation, keep the animals in a warm cage as long as no bleeding or tumor tissue leakage occurs. Monitor the animal until they regain sufficient consciousness to maintain sternal recumbency. Return the animal to the animal room after full recovery from anesthesia. Monitor the tumor-bearing mice by palpating the abdomen near the spleen and select out the mice bearing orthotopic tumors.