Ovarian Carcinoma Tubule Formation Assay

This procedure was previously described in [25 (link)]. Human ovarian carcinoma cell SKOV3 and human adherent ovarian carcinoma cells were seeded on Matrigel at a density of 10 ⁵ cells per well in complete specific medium incubated at 37 °C with 5% CO₂, and were then observed for their capacity to form tubule-like structures. ED-B was added at a concentration of 2 g/mL after two hours when ovarian carcinoma cells started to form tubules, to test the ability of the ED-B fragment of fibronectin to induce tubule formation via SKOV3 and ovarian carcinoma cell OS13. To test the ability of OC-46F2 and anti-B-FN scFv to inhibit tubule formation via SKOV3 and OS13, antibodies were added at a concentration of 200 g/mL when ovarian carcinoma cells started to form tubules. All experiments were performed in triplicate. After fixation in 2% paraformaldehyde in PBS, the tubules formed were counted on ten different high magnification microscopic fields per coverslip under light microscopy (Leica Microsystems, Wetzlar, Germany) at 100X. Images were captured using a DM LB2 microscope camera (Leica) [53 (link)].