Cell invasion rates were established using our previously published method
[18 (link),19 (link)]. Using a real-time cell imaging system (IncuCyte™ (Essen BioScience, Michigan, USA). In brief, 96-well plates were coated with a thin layer of collagen by transferring 300 μg/ml of collagen type I (Life Technologies™, Carlsbad, CA) and incubating at 37°C for 30 min. OVCAR-3 and SKOV-3 (2 × 104 cells per well) were grown to confluence in complete growth media. Cell-free zones were created by generating a wound with a 96-Well WoundMaker™ (Essen BioScience, Michigan, USA). The cells were overlaid with 3 mg/ml collagen type I (Life Technologies™, Carlsbad, CA) and incubated at 37°C for 30 min to create a 3D matrix. Complete growth media was added on top of the layer of collagen. Cells were imaged automatically every 3 h over a time period of 48 h. The images were processed by the IncuCyte™ software package (Essen BioScience, Michigan, USA) to measure cell invasion by obtaining the Relative Wound Density (RWD, as defined by custom algorithms within the IncuCyte™ software package). These users informed algorithms identify the wound region and provide visual representations of the segmentation parameters. Image collection was created using three to five representative phase contrast images.
[18 (link),19 (link)]. Using a real-time cell imaging system (IncuCyte™ (Essen BioScience, Michigan, USA). In brief, 96-well plates were coated with a thin layer of collagen by transferring 300 μg/ml of collagen type I (Life Technologies™, Carlsbad, CA) and incubating at 37°C for 30 min. OVCAR-3 and SKOV-3 (2 × 104 cells per well) were grown to confluence in complete growth media. Cell-free zones were created by generating a wound with a 96-Well WoundMaker™ (Essen BioScience, Michigan, USA). The cells were overlaid with 3 mg/ml collagen type I (Life Technologies™, Carlsbad, CA) and incubated at 37°C for 30 min to create a 3D matrix. Complete growth media was added on top of the layer of collagen. Cells were imaged automatically every 3 h over a time period of 48 h. The images were processed by the IncuCyte™ software package (Essen BioScience, Michigan, USA) to measure cell invasion by obtaining the Relative Wound Density (RWD, as defined by custom algorithms within the IncuCyte™ software package). These users informed algorithms identify the wound region and provide visual representations of the segmentation parameters. Image collection was created using three to five representative phase contrast images.
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