Rtci precoated

An organotypic mesomimetic model was utilized to monitor cell-to-mesothelium adhesion [40] (link). OvCa433Ncad+ and SKOV3Ecad+ cells were transiently stained with red CMTPX or green CMFDA (Invitrogen), respectively. Parental OvCa433 and SKOV3 cells were RFP- or GFP-tagged, respectively, as described. Cells were applied to LP9 human mesothelial monolayers in RTCI-precoated (Corning) 24-well dishes at 200,000 cells/ml (1ml total), incubated for 30 or 60 minutes, washed 3× with PBS washes, and imaged with AMG EVOS fluorescence microscope. Image analysis was performed using ImageJ. The assay was repeated in triplicate, and statistical analysis was performed using a Student's t test.

Cells were diluted to 100,000 cells/ml and 500 μl seeded on top of RTCI-precoated (10 μg/ml in 0.1 M sodium carbonate pH 9.6; Corning) 24-well dishes and incubated for 10 to 60 minutes, then fixed and stained with Diff-Quick Kit (Siemens) and adherent cells enumerated. The assay was repeated in triplicate, and statistical analysis was performed using a Student's t test.