Modeling Ovarian Cancer Cell Colonization of Omentum: An Ex Vivo Method to Establish Milky Spot Colonization of Fluorescently Labeled Ovarian Cancer Cells in Murine Omentum Explants

Grow and prepare fluorescently tagged cells and resuspend at a concentration of 2 million cells per milliliter. Apply approximately 6 microliters of the tissue adhesive to the membrane of the culture insert and allow it to air dry. Then, wash the membrane twice with sterile water to remove any excessive adhesive before air drying the membranes under a laminar hood.

Carefully excise the omenta and attach it to the adhesive-coated membrane using sterile forceps. After allowing the tissue to adhere to the membrane for one minute, add 500 microliters of the cell suspension on top of each omentum in each culture insert.

Then, fill the area around the transwell chamber with 2.5 milliliters of DME/F-12 media. Incubate the omenta with cell suspension for six hours at 37 degrees Celsius in a 5% CO2 environment. Carefully remove and wash the omenta with about 10 milliliters of PBS. Finally, visualize fluorescent cancer cell foci using an appropriate fluorescent imaging system.