Fig. 7.

Effects of Cal33 cell number on fluorescent drug distribution in 2D monolayers: images and pseudo-color pixel intensity visualizations for (A) test compounds at a seeding density of 10,000 cells/well, (B) ellipticine, and (C) doxorubicin at different seeding densities. Cal33 cells were seeded into 384-well monolayer assay plates at seeding densities of 625, 1,250, 2,500, 5,000, 10,000, and 20,000 cells per well, and after 24 h in culture, 10 μM of test compounds was added to wells and incubated for 15 min before being fixed in formaldehyde and washed with PBS. FITC and TRITC images of Cal33 monolayers were acquired on the IXM. Representative images colored by the image acquisition channel, green for FITC and yellow for TRITC, are presented along with the corresponding pseudo-color pixel intensity visualizations, where the relative fluorescent intensities of the pixels in the images are indicated as distinct colors. The “hotter” and “brighter” colors (low to high, yellow, red, white) represent higher intensity signals, and cooler colors (low to high, purple, cyan, green) represent lower intensity signals. Representative data from one of three independent experiments are shown.