This article details the procedure for orthotopic inoculation of ovarian cancer cells in a nude mouse model. The method allows for the study of tumor growth and metastasis in a controlled environment.
Orthotopic inoculation helps deliver cancer cells to a recipient animal at a specific tissue or site matching the cells' natural anatomical location. To begin, prep an anesthetized female nude mouse, belly side down, on an operating stage. Make a vertical incision near its spine at a position between the rib cage and the femur. Open the abdominal muscles to expose the peritoneal cavity partially. Locate one of the ovaries.
Prepare a suspension of ovarian cancer cells with chilled extracellular matrix in a syringe. The low temperature prevents the suspension from solidifying. Inject this suspension into the mouse's ovary. Hold the needle in place to allow the matrix to solidify and embed the cancer cells within the ovary. Retract the needle. Suture the incision to close the wound. Allow the mouse to recover.
Over time, the injected ovarian cancer cells proliferate to form a primary tumor. Eventually, a few cells detach from their primary location and disseminate within the peritoneal cavity. When these migrating tumor cells attach to the peritoneal cavity lining, they invade the peritoneal tissue and proliferate to form metastasized secondary tumors.
To carry out orthotopic inoculation, after anesthetizing an eight-week-old female nude mouse according to the text protocol, perform a toe pinch to verify depth of the anesthesia. Place the anesthetized mouse on an operating stage, belly side down. Then, disinfect the surgical area with several washes using a chlorhexidine-based scrub and alcohol.
Next, make an approximately 2-centimeter vertical incision near the spine, between the right costal arch and the femur. Place a sterile surgical drape around the incision site. Then, fix the incised skin surfaces with clamps to maintain the opening.
Now, make a second 1-centimeter incision at the parietal peritoneum under the first incision to open the abdominal cavity. Then, clamp the incised abdominal surfaces. With a clamp, clip the fat tissue that surrounds the ipsilateral ovary and fallopian tube to remove the ovary from the abdominal cavity.
Then, carefully place the ovary and fallopian tube on gauze before placing the mouse, belly side down, under the dissecting microscope. Transfer the cell suspension to an insulin syringe. Then, immediately and carefully inject 1 to 2 microliters of the prepared cell suspension into the ovary.
For several seconds, keep the needle in the ovary to ensure that the hydrogel sets. After gelation, the injected tumor cells will remain in the ovary. Carefully, return the ovary into the body. With sterile medical silk, suture the second incision before using wound clips to seal the skin.
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