Mesothelial cells were cultured on 6-well plastic plates (Techno Plastic Products) and used in the clearance assay once a confluent monolayer was formed (approximately 72 hours). Mesothelial cells were labeled 18 hours prior with CMFDA-green (Molecular Probes), washed with PBS and incubated with fresh culture medium until use. Spheroids were prepared 18 hours in advance. Cells were labeled with CMTPX-red (Molecular Probes), washed with PBS, dissociated by trypsinization and resuspended in culture medium. Labeled cells were then counted and plated at 200 cells/well in ultra-low attachment multiwell plates (Corning) with 10ug/mL of soluble bovine fibronectin for increased cohesion. Spheroids were placed onto the monolayer of mesothelial cells at the microscope and images captured at 0, 1, and 7 hours. Experiment replicated 3 independent times. Normalized clearance value is determined by measuring the total mesothelial cell area cleared by the spheroid in ImageJ, and normalizing to the size of the spheroid at time 0.
6 well plastic plates
Manufactured by Techno Plastic Products
6-well plastic plates are a laboratory equipment used to hold and culture cells or tissues in a controlled environment. Each plate contains six individual wells, providing a convenient and standardized platform for various cell-based experiments and assays.
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2 protocols using 6 well plastic plates
1
Mesothelial Cell Clearance Assay
2
Mesothelial Cell Clearance Assay
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Mesothelial cells were cultured on 6-well plastic plates (Techno Plastic Products) and used in the clearance assay once a confluent monolayer was formed (approximately 72 hours). Mesothelial cells were labeled 18 hours prior with CMFDA-green (Molecular Probes), washed with PBS and incubated with fresh culture medium until use. Spheroids were prepared 18 hours in advance. Cells were labeled with CMTPX-red (Molecular Probes), washed with PBS, dissociated by trypsinization and resuspended in culture medium. Labeled cells were then counted and plated at 200 cells/well in ultra-low attachment multiwell plates (Corning) with 10ug/mL of soluble bovine fibronectin for increased cohesion. Spheroids were placed onto the monolayer of mesothelial cells at the microscope and images captured at 0, 1, and 7 hours. Experiment replicated 3 independent times. Normalized clearance value is determined by measuring the total mesothelial cell area cleared by the spheroid in ImageJ, and normalizing to the size of the spheroid at time 0.
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