Pre-coat the entire well surface of the CytoView-Z plate using an extracellular matrix molecule (ECMM), such as fibronectin.
Incubate the surface-coated plate in a cell culture incubator at 37°C and 5% CO2 for at least 1 hour.
Aspirate the surface coating solution from the plate.
Add 100 µl of complete medium to the plate, and add 8 mL of sterile water to the onplate reservoirs to increase humidity.
Dock the plate in the Maestro Z and measure the media only (MO) baseline. Transfer the plate to a biosafety cabinet when the baseline is complete.
Culturing Adherent Target Cells and Transferring to CytoView-Z Plate
Thaw and culture the target cells of interest in accordance with supplier recommendations, passaging as needed.
Remove flasks of cultured cells from the incubator, aspirate the media, and rinse with warmed phosphate-buffered saline (PBS). Using trypsin, or another cell dissociating agent, detach and collect the cells from the flasks as per reagent recommendations.
Remove a sample of the cell suspension and count the cells using the Exact FL or a hemocytometer to determine both the cell viability and total number of viable cells.
Transfer the cell suspension to a 15 ml conical tube and centrifuge the cell suspension to pellet.
Aspirate the supernatant, being careful not to disturb the cell pellet.
Dilute the cell suspension in complete medium to the desired working concentration of cells per 100 µl.
Plating Adherent Target Cells onto the CytoView-Z Plate
Transfer the cell suspension to a reservoir for easy access by a multichannel pipette. Add 100 µl of cell suspension to the 100 µl of media already present in each well.
Leave the plate seeded with target cells to rest in the biosafety cabinet for 1 hour at room temperature.
Dock the plate and impedance measurements will begin automatically upon plate engagement.
Dosing Target Cancer Cells with Effector Cells in the CytoView-Z Plate
Thaw and culture the cells of interest in accordance with supplier recommendations.
At 24 hrs following target cell seeding, take the flasks of cultured effector cells from the 37°C incubator and mix the suspension gently but thoroughly with a serological pipettor.
Remove a sample of the cell suspension and count the cells using the Exact FL or a hemocytometer to determine both the viability and total number of viable cells.
Transfer the cell suspension to a 15 ml conical tube and centrifuge the cell suspension to pellet.
Aspirate the supernatant, being careful not to disturb the cell pellet.
Dilute the cell suspension in complete effector cell medium to the desired working concentration of cells per 22.2 µl and add 22.2 µl of effector cell suspension to each well.
Leave the plate dosed with effector cells to rest in the biosafety cabinet for 30 minutes at room temperature.
Dock the plate and impedance measurements will automatically resume upon plate engagement.
