Intraperitoneal Injection of Cells: A Method of Delivering Cancer Cells into the Peritoneal Adipose Tissue of Murine Model

Overview

In this video, we perform an intraperitoneal injection of ovarian cancer cell suspension in a mouse model to study the colonization of ovarian cancer cells in peritoneal adipose tissues and their subsequent metastasis. 

Protocol

All procedures involving animals have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.

1. Intraperitoneal Injection of Cells

NOTE: In our experiments, mice are handled in a laminar airflow or biosafety cabinet within our barrier facility in order to limit the risk of pathogen exposure. In order to demonstrate this technique clearly, the procedures in the accompanying video were conducted in a laboratory approved for animal work. This particular technique does not require the animals to be under anesthesia. Under approved protocols, perform this technique on live animals. Use appropriate strains of mice for this study. For example: use immunocompromised, athymic nude mice for the study of human ovarian cancer cell (SKOV3ip.1 and HeyA8) colonization; use immunocompetent C57BL/6 mice for study of mouse ovarian cancer cell (ID8) colonization.

1. Load 500 µl of single cell suspension of ovarian cancer cells into a syringe and put in the sterile capped 25 G needle. This reduces cell shearing prior to injection.
2. Pick the mouse up by the scruff of the neck and hold the tail using the palm and forefinger and fix the left hind leg between the ring and little finger (when the mouse is restrained with the left hand).
   NOTE: To avoid traumatizing abdominal organs, restrain the mouse well so that it cannot move during the injection.
3. Imagine a line across the abdomen just above the knees, and locate a point on the animal's right side and close to the midline. The point of entry is cranial to and slightly medial of the last nipple.
   NOTE: Inserting the needle on the mouse's right side avoids the cecum and reduces the risk of puncturing the intestines.
4. Insert the needle at the lower lateral region of the mice's abdomen to a depth of approximately 0.5 cm. Pull back on the plunger to confirm that the needle has not penetrated a blood vessel or other peritoneal organs.
   1. If any fluid is aspirated discard the syringe (and sample). A greenish-brown or yellow aspirate indicates needle penetration into the intestines or bladder, respectively.
5. Inject the sample using slow, steady pressure. Withdraw the needle and return the mouse to its cage. Do not recap the syringe before disposal in a sharp container.
6. Sacrifice mice via CO₂ asphyxiation and vital organ removal at a specific time point post injection.

Materials

Name	Company	Catalog Number	Comments
DMEM	Corning	10-013-CV
PBS	Corning	21-040-CV	Without calcium and Magnesium
26 gauge needle	BD	329652